Fungal Defensin Research Articles

Fungal Defensins: Dual Roles in Itch and Inflammation

Fungal Defensins: Dual Roles in Itch and Inflammation

Characterization and expression of fungal defensin in Escherichia coli and its antifungal mechanism by RNA-seq analysis.

Invasive fungal infections (IFIs) are fatally threatening to critical patients. The fungal defensin as an antifungal protein can widely inhibit fungi. In this study, eight antifungal genes from different filamentous fungi were optimized by synonymous codon bias and heterologously expressed in Escherichia coli. Only the antifungal protein (AFP) from Aspergillus giganteus was produced, whereas the AFP from its mutation of the chitin-binding domain could not be expressed, thereby suggesting the importance of the motif for protein folding. In addition, the recombinant AFP (rAFP, 100 μg/mL) pre-heated at 50°C for 1 h effectively inhibited Paecilomyces variotii CICC40716 of IFIs by 55%, and no cell cytotoxicity was observed in RAW264.7 cells. After being pre-heated at 50°C for 8 h, the fluorescence emission intensity of the rAFP decreased and shifted from 343 nm to 335 nm. Moreover, the helix and β-turn of the rAFP gradually decreased with the pre-heated treatment temperature of 50°C via circular dichroism spectroscopy. Propidium iodide staining revealed that the rAFP could cause damage to the cell membrane. Moreover, the corresponding differentially expressed genes (DEGs) for downregulation such as amino sugar and nucleotide sugar metabolism, as well as mitogen-activated protein kinase (MAPK) signaling pathway involved in the cell wall integrity were found via the RNA-seq of rAFP treatment. By contrast, the upregulated DEGs were enriched in response to the oxidative stress of Biological Process by the Gene Ontology (GO) database. The encoding proteins of laccase, multicopper oxidase, and nitroreductase that contributed to reactive oxygen species (ROS) scavenging could be recognized. These results suggested that the rAFP may affect the integrity of the cell wall and cell membrane, and promote the increase in ROS, thereby resulting in fungal death. Consequently, drug development could be based on the inhibitory effect of the rAFP on IFIs.

Molecular Modification of Kex2 P1' Site Enhances Expression and Druggability of Fungal Defensin.

Pichia pastoris is the widely used expression system for producing recombinant secretory proteins. It is known that Kex2 protease plays a vital role in the process of protein secretion, in which the P1' site affects its cleavage efficiency. To enhance the expression level of fungal defensin-derived peptide NZ2114, this work attempts to optimize the P1' site of Kex2 by replacing it with 20 amino acids in turn. The results showed that when the amino acid of the P1' site was changed to Phe (F), the yield of target peptide significantly increased from 2.39 g/L to 4.81 g/L. Additionally, the novel peptide F-NZ2114 (short for FNZ) showed strong antimicrobial activity against Gram-positive (G+) bacteria, especially for Staphylococcus aureus and Streptococcus agalactiae (MIC: 4-8 μg/mL). The FNZ was very stable and retained high activity in various conditions; in addition, a low cytotoxicity and no hemolysis were observed even at a high concentration of 128 μg/mL, and a longer postantibiotic effect was reached. The above results indicate that this engineering strategy provided a feasible optimization scheme for enhancing the expression level and druggability of this antimicrobial peptide from fungal defensin and other similar targets by this updated recombinant yeast.

Suppression of the insect cuticular microbiomes by a fungal defensin to facilitate parasite infection.

Insects can assemble defensive microbiomes on their body surfaces to defend against fungal parasitic infections. The strategies employed by fungal pathogens to combat host cuticular microbiotas remains unclear. Here, we report the identification and functional characterization of the defensin-like antimicrobial gene BbAMP1 encoded by the entomopathogenic fungus Beauveria bassiana. The mature peptide of BbAMP1 can coat fungal spores and can be secreted by the fungus to target and damage Gram-positive bacterial cells. Significant differences in insect survival were observed between the wild-type and BbAMP1 mutant strains during topical infection but not during injection assays that bypassed insect cuticles. Thus, BbAMP1 deletion considerably reduced fungal virulence while gene overexpression accelerated the fungal colonization of insects compared with the wild-type strain in natural infections. Topical infection of axenic Drosophila adults evidenced no difference in fly survivals between strains. However, the gnotobiotic infections with the addition of Gram-positive but not Gram-negative bacterial cells in fungal spore suspensions substantially increased the survival of the flies treated with ∆BbAMP1 compared to those infected by the wild-type and gene-overexpression strains. Bacterial colony counts and microbiome analysis confirmed that BbAMP1 could assist the fungus to manipulate insect surface bacterial loads. This study reveals that fungal defensin can suppress the host surface defensive microbiomes, which underscores the importance to extend the research scope of fungus-host interactions.

Effect of NZ2114 against Streptococcus dysgalactiae biofilms and its application in murine mastitis model.

Bovine mastitis caused by Streptococcus dysgalactiae (S. dysgalactiae) is usually treated with antibiotics, which may potentially increase drug resistance as the abuse. NZ2114, a variant of fungal defensin plectasin, displayed a potent antibacterial activity against S. dysgalactiae. The inhibition/eradication effect of the antimicrobial peptide NZ2114 on the early/mature biofilm of S. dysgalactiae CVCC 3938 was evaluated, as well as the elimination of bacteria in mature biofilms. In this study, NZ2114 displayed potent antibacterial activity against S. dysgalactiae CVCC 3938 and three clinical isolated S. dysgalactiae strains (0.11-0.45 μM). The early biofilm inhibition of S. dysgalactiae CVCC 3938 was 55.5–85.9% after treatment with NZ2114 at concentrations of 1–16 × MIC, which was better than that of vancomycin at the same concentration. The mature biofilm eradication rate was up to 92.7–97.6% with the increasing concentration (2–16 × MIC) of NZ2114, and the eradication rate did not change significantly with further increase of NZ2114 concentration, while the biofilm eradication rate of vancomycin-treated group at the same concentration remained at 92.5%. NZ2114 reduced the number of persister bacteria in biofilm. Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) further demonstrated that NZ2114 could effectively reduce the biofilm thickness and bacterial number of S. dysgalactiae CVCC 3938. In vivo therapeutic effect of NZ2114 on murine mastitis model showed that NZ2114 was better than vancomycin in alleviating mammary gland inflammation by regulating cytokines production, inhibiting bacterial proliferation, and reducing the number of mammary gland bacteria. These data suggested that NZ2114 is a potential peptide candidate for the treatment of mastitis.

Pharmacokinetics and Pharmacodynamics of Fungal Defensin NZX Against Staphylococcus aureus-Induced Mouse Peritonitis Model.

Staphylococcus aureus (S. aureus) is one of the most common pathogenic bacteria responsible for causing a life-threatening peritonitis disease. NZX, as a variant of fungal defensin plectasin, displayed potent antibacterial activity against S. aureus. In this study, the antibacterial and resistance characteristics, pharmacokinetics, and pharmacodynamics of NZX against the S. aureus E48 and S. aureus E48-induced mouse peritonitis model were studied, respectively. NZX exhibited a more rapid killing activity to S. aureus (minimal inhibitory concentration, 1 μg/ml) compared with linezolid, ampicillin and daptomycin, and serial passaging of S. aureus E48 for 30 days at 1/2 × MIC, NZX had a lower risk of resistance compared with ampicillin and daptomycin. Also, it displayed a high biocompatibility and tolerance to physiological salt, serum environment, and phagolysosome proteinase environment, except for acid environment in phagolysosome. The murine serum protein-binding rate of NZX was 89.25% measured by ultrafiltration method. Based on the free NZX concentration in serum after tail vein administration, the main pharmacokinetic parameters for T1/2, Cmax, Vd, MRT, and AUC ranged from 0.32 to 0.45 h, 2.85 to 20.55 μg/ml, 1469.10 to 2073.90 ml/kg, 0.32 to 0.56 h, and 1.11 to 8.89 μg.h/ml, respectively. Additionally, the in vivo pharmacodynamics against S. aureus demonstrated that NZX administrated two times by tail vein at 20 mg/kg could rescue all infected mice in the lethal mouse peritonitis model. And NZX treatment (20 mg/kg) significantly reduced CFU counts in the liver, lung, and spleen, especially for intracellular bacteria in the peritoneal fluid, which were similar or superior to those of daptomycin. In vivo efficacies of NZX against total bacteria and intracellular bacteria were significantly correlated with three PK/PD indices of ƒAUC/MIC, ƒCmax/MIC, and ƒT% > MIC analyzed by a sigmoid maximum-effect model. These results showed that NZX may be a potential candidate for treating peritonitis disease caused by intracellular S. aureus.

A Fungal Defensin Inhibiting Bacterial Cell-Wall Biosynthesis with Non-Hemolysis and Serum Stability.

Defensins are a class of cationic disulfide-bridged antimicrobial peptides (AMPs) present in many eukaryotic organisms and even in bacteria. They primarily include two distinct but evolutionarily related superfamilies (cis and trans). Defensins in fungi belong to the members of the cis-superfamily with the cysteine-stabilized α-helical and β-sheet fold. To date, many fungal defensin-like peptides (fDLPs) have been found through gene mining of the genome resource, but only a few have been experimentally characterized. Here, we report the structural and functional characterization of Pyronesin4 (abbreviated as Py4), a fDLP previously identified by genomic sequencing of the basal filamentous ascomycete Pyronema confluens. Chemically, synthetic Py4 adopts a native-like structure and exhibits activity on an array of Gram-positive bacteria including some clinical isolates of Staphylococcus and Staphylococcus warneri, a conditioned pathogen inhabiting in human skin. Py4 markedly altered the bacterial morphology and caused cytoplasmic accumulation of the cell-wall synthesis precursor through binding to the membrane-bound Lipid II, indicating that it works as an inhibitor of cell-wall biosynthesis. Py4 showed no hemolysis and high mammalian serum stability. This work identified a new fungal defensin with properties relevant to drug exploration. Intramolecular epistasis between mutational sites of fDLPs is also discussed.

A study on fungal defensin against multidrug-resistant Clostridium perfringens and its treatment on infected poultry.

In the present study, we aimed to investigate the antibacterial activity and mechanisms of plectasin-derived peptide NZ2114 in vitro and its therapeutic effects in vivo on broilers challenged with Clostridium perfringens. In vitro assay showed that NZ2114 had potent (minimal inhibitory concentration, 0.91μM) and rapid antibacterial activity (99.9% reduction within 2h), and the dual antibacterial mechanisms (including interfering with the cell membrane and intracellular DNA) against C. perfringens CVCC 2030. In vivo study, NZ2114 tended to increase linearly and quadratically the average daily gain as NZ2114 level increased and was the highest at 20mg/L. NZ2114 at 10 ~ 40mg/L dramatically reduced jejunal lesion score. Besides, the levels of IL-6, TNF-α, and IL-1β tended to downregulate linearly and quadratically as the NZ2114 level increased and were all the lowest at the dose of 20mg/L. NZ2114 significantly upregulated those levels of IgA, IgG, IgM, and sIgA with a linear and quadratic dose effect, with the highest IgA, IgG, IgM, and sIgA at 20mg/L. Finally, NZ2114 tended to linearly and quadratically increase the numerical value of crypt depth, with the lowest value at 40mg/L. Lincomycin only dramatically reduced the jejunal lesion score and increased the numerical value of crypt depth. These results indicate that NZ2114 has the potential as a new alternative to antibiotics for the treatment of C. perfringens-induced necrotic enteritis infection.Key points• NZ2114 could kill C. perfringens by dual antibacterial mechanisms• Broiler necrotic enteritis model induced by C. perfringens was established• NZ2114 treatment could ameliorate C. perfringens-induced necrotic enteritis.

A Fungal Defensin Targets the SARS-CoV-2 Spike Receptor-Binding Domain.

Coronavirus Disease 2019 (COVID−19) elicited by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS−CoV−2) is calling for novel targeted drugs. Since the viral entry into host cells depends on specific interactions between the receptor−binding domain (RBD) of the viral Spike protein and the membrane−bound monocarboxypeptidase angiotensin converting enzyme 2 (ACE2), the development of high affinity RBD binders to compete with human ACE2 represents a promising strategy for the design of therapeutics to prevent viral entry. Here, we report the discovery of such a binder and its improvement via a combination of computational and experimental approaches. The binder micasin, a known fungal defensin from the dermatophytic fungus Microsporum canis with antibacterial activity, can dock to the crevice formed by the receptor−binding motif (RBM) of RBD via an extensive shape complementarity interface (855.9 Å2 in area) with numerous hydrophobic and hydrogen−bonding interactions. Using microscale thermophoresis (MST) technique, we confirmed that micasin and its C−terminal γ−core derivative with multiple predicted interacting residues exhibited a low micromolar affinity to RBD. Expanding the interface area of micasin through a single point mutation to 970.5 Å2 accompanying an enhanced hydrogen bond network significantly improved its binding affinity by six−fold. Our work highlights the naturally occurring fungal defensins as an emerging resource that may be suitable for the development into antiviral agents for COVID−19.

A recombinant fungal defensin-like peptide-P2 combats Streptococcus dysgalactiae and biofilms.

Streptococcus dysgalactiae, considered one of the main pathogens that causes bovine mastitis, is a serious threat to humans and animals. However, the excessive use of antibiotics and the characteristic of S. dysgalactiae forming biofilms in mastitic teat canal have serious clinical implications. In this study, in vivo and in vitro multiple mechanisms of action of P2, a mutant of fungal defensin plectasin, against S. dysgalactiae were systematically and comprehensively investigated for the first time. P2 showed potent antibacterial activity against S. dysgalactiae (minimum inhibitory concentration, MIC = 0.23-0.46 μM) and rapid bactericidal action by 3.0 lg units reduction in 2-4 h. No resistant mutants appeared after 30-d serial passage of S. dysgalactiae in the presence of P2. The results of electron microscopy and flow cytometer showed that P2 induced membrane damage of S. dysgalactiae, causing the leakage of cellular content and eventually cell death. Besides, P2 effectively inhibited early biofilm formation, eradicated mature biofilms, and killed 99.9% persisters which were resistant to 100 × MIC vancomycin; and confocal laser scanning microscopy (CLSM) also revealed the potent antibacterial and antibiofilm activity of P2 (the thickness of biofilm reduced from 18.82 to 7.94 μm). The in vivo therapeutic effect of P2 in mouse mastitis model showed that it decreased the number of mammary bacteria and alleviated breast inflammation by regulating cytokines and inhibiting bacterial proliferation, which were superior to vancomycin. These data indicated that P2 maybe a potential candidate peptide for mastitis treatment of S. dysgalactiae infections. KEY POINTS: •P2 showed potential in vitro antibacterial characteristics towards S. dysgalactiae. •P2 eradicated biofilms, killed persisters, and induced cell death of S. dysgalactiae. •P2 could effectively protect mice from S. dysgalactiae infection in gland.

The second N-terminal aromatic residue of the fungal defensin, blapersin, of Blastomyces percursus is essential for its antibacterial activity

Studies have shown that the second N-terminal residue of fungal defensins is closely involved in the binding of defensins to lipid II, a bacterial cell wall precursor, and plays an important role in antibacterial activity. We found that the N-terminal residue is always aromatic in nature. In this study, 29 fungal defensin-like peptides were found via the genomic search strategy. Based on the type of aromatic residue at the second N-terminal site, we mainly divided these peptides into Phe, Trp, and Tyr types. We selected and characterized a defensin, blapersin, derived from Blastomyces percursus as a molecular model to investigate the functional significance of the N-terminal site. The native blapersin killed a wide spectrum of gram-positive bacteria at low molecular concentrations. Its aromatic mutants, W2F and W2Y, displayed enhanced antimicrobial activity, especially against the vancomycin-resistant Enterococcus faecium. The aromatic side chains containing Phe2 and Tyr2 seem to be more favorable for the antibacterial activity of blapersin those containing Trp2. However, the nonaromatic mutant W2A had almost no antibacterial activity. This indicates that the second N-terminal aromatic residue is essential for the antimicrobial action of blapersin. All these defensins have high stability and low toxicity. This is the first report on the enhancement of antibacterial activity by calibration of the N-terminal aromatic residue.

Bioactive Peptides Against Fungal Biofilms.

Infections caused by invasive fungal biofilms have been widely associated with high morbidity and mortality rates, mainly due to the advent of antibiotic resistance. Moreover, fungal biofilms impose an additional challenge, leading to multidrug resistance. This fact, along with the contamination of medical devices and the limited number of effective antifungal agents available on the market, demonstrates the importance of finding novel drug candidates targeting pathogenic fungal cells and biofilms. In this context, an alternative strategy is the use of antifungal peptides (AFPs) against fungal biofilms. AFPs are considered a group of bioactive molecules with broad-spectrum activities and multiple mechanisms of action that have been widely used as template molecules for drug design strategies aiming at greater specificity and biological efficacy. Among the AFP classes most studied in the context of fungal biofilms, defensins, cathelicidins and histatins have been described. AFPs can also act by preventing the formation of fungal biofilms and eradicating preformed biofilms through mechanisms associated with cell wall perturbation, inhibition of planktonic fungal cells’ adhesion onto surfaces, gene regulation and generation of reactive oxygen species (ROS). Thus, considering the critical scenario imposed by fungal biofilms and associated infections and the application of AFPs as a possible treatment, this review will focus on the most effective AFPs described to date, with a core focus on antibiofilm peptides, as well as their efficacy in vivo, application on surfaces and proposed mechanisms of action.

Oxidative stress and the presence of bacteria increase gene expression of the antimicrobial peptide aclasin, a fungal CSαβ defensin in Aspergillus clavatus.

BackgroundAntimicrobial peptides (AMPs) represent a broad class of naturally occurring antimicrobial compounds. Plants, invertebrates and fungi produce various AMPs as, for example, defensins. Most of these defensins are characterised by the presence of a cysteine-stabilised α-helical and β-sheet (CSαβ) motif. The changes in gene expression of a fungal CSαβ defensin by stress conditions were investigated in Aspergillus clavatus. A. clavatus produces the CSαβ defensin Aclasin, which is encoded by the aclasin gene.MethodsAclasin expression was evaluated in submerged mycelium cultures under heat shock, osmotic stress, oxidative stress and the presence of bacteria by quantitative real-time PCR.ResultsAclasin expression increased two fold under oxidative stress conditions and in the presence of viable and heat-killed Bacillus megaterium. Under heat shock and osmotic stress, aclasin expression decreased.DiscussionThe results suggest that oxidative stress and the presence of bacteria might regulate fungal defensin expression. Moreover, fungi might recognise microorganisms as plants and animals do.

Recombinant AfusinC, an anionic fungal CSαβ defensin from Aspergillus fumigatus, exhibits antimicrobial activity against gram-positive bacteria.

Antimicrobial peptides (AMPs) are short and generally positively charged peptides found in a wide variety of organisms. CSαβ defensins are a group of AMPs. These defensins are composed of an α-helix and a β-sheet linked by three or four disulphide bridges. In this study, we describe the antimicrobial activity of an anionic CSαβ fungal defensin from Aspergillus fumigatus, AfusinC. AfusinC was recombinantly produced as a fusion protein in Escherichia coli. The tag was removed by proteolytic cleavage, and AfusinC was purified by size exclusion chromatography. About 0.8 mg of recombinant AfusinC was obtained from 1 L of culture. Recombinant AfusinC was active against mainly gram-positive bacteria including human pathogens and a multiresistant-strain of A. aureus. Additionally, AfusinC showed bactericidal effect against Micrococcus luteus.

Triintsin, a human pathogenic fungus-derived defensin with broad-spectrum antimicrobial activity

Since there is a symbiotic and competitive relationship between microorganisms in the same ecological niche, fungal defensins have been found to be important resources for antimicrobial peptides. Here, a fungal defensin, triintsin, was characterized in a clinical isolate of Trichophyton interdigitale from a patient with onychomycosis. The comparison of its genomic and mRNA sequences showed the gene organization and structure of three coding exons separated by two introns. The precursor peptide of triintsin contained 85 amino acid residues, which were composed of three parts including an N-terminal signal domain of 21 residues, a pro-peptide of 47 residues that ended at lysine-arginine and a mature peptide of 38 residues at the C-terminus. The 3D-structure established by homology modeling revealed that triintsin presented a representative typical cysteine-stabilized α-helical and β-sheet fold. The reductive linear peptide of triintsin was obtained by chemical synthesis. After cyclization to form three pairs of disulfide bonds, the oxidative-type peptide displayed broad-spectrum antimicrobial activity against both gram-positive and gram-negative bacteria but also showed anti-fungal activity. Moreover, triintsin can effectively inhibit the growth of clinical strains. Altogether, the peptide is a human pathogenic fungus-derived defensin with broad-spectrum antimicrobial activity.

Single-point mutation-mediated local amphipathic adjustment dramatically enhances antibacterial activity of a fungal defensin.

The emergence and rapid spread of multiresistant bacteria has lead to an urgent need for novel antimicrobials. Based on single-point substitutions, we generated a series of mutants of micasin, a dermatophytic defensin, with enhanced activities against multiple clinical isolates of Staphylococcus species, including 4 antibiotic-resistant strains. We first mapped the functional surface of micasin by alanine-scanning mutational analysis of its highly exposed residues, through which we found that substitution of site 8 (acidic Glu) dramatically enhanced bacterial killing of this peptide. Structural analysis indicates that this single point mutation could result in a functional local amphipathic architecture. Four different types of side chains (hydrophobic, cationic polar, neutral polar, and acidic polar) were introduced at site 8 to clarify the role of this local architecture in micasin function. The results show that all mutants displayed increased antibacterial activity with the exception of the acidic replacement. These mutants with enhanced activity exhibited low hemolysis and cytotoxicity and showed high serum stability, indicating their therapeutic potential. Our work represents the first example of structural fine-tuning to largely improve the antibacterial potency of a dermatophytic defensin.-Wu, J., Gao, B., Zhu, S. Single-point mutation-mediated local amphipathic adjustment dramatically enhances antibacterial activity of a fungal defensin.

Cysteine-stabilized αβ defensins: From a common fold to antibacterial activity

Antimicrobial peptides (AMPs) seem to be promising alternatives to common antibiotics, which are facing increasing bacterial resistance. Among them are the cysteine-stabilized αβ defensins. These peptides are small, with a length ranging from 34 to 54 amino acid residues, cysteine-rich and extremely stable, normally composed of an α-helix and three β-strands stabilized by three or four disulfide bonds and commonly found in several organisms. Moreover, animal and plant CSαβ defensins present different specificities, the first being mainly active against bacteria and the second against fungi. The role of the CSαβ-motif remains unknown, but a common antibacterial mechanism of action, based on the inhibition of the cell-wall formation, has already been observed in some fungal and invertebrate defensins. In this context, the present work aims to group the data about CSαβ defensins, highlighting their evolution, conservation, structural characteristics, antibacterial activity and biotechnological perspectives.

Plectasin, first animal toxin-like fungal defensin blocking potassium channels through recognizing channel pore region.

The potassium channels were recently found to be inhibited by animal toxin-like human β-defensin 2 (hBD2), the first defensin blocker of potassium channels. Whether there are other defensin blockers from different organisms remains an open question. Here, we reported the potassium channel-blocking plectasin, the first defensin blocker from a fungus. Based on the similar cysteine-stabilized alpha-beta (CSαβ) structure between plectasin and scorpion toxins acting on potassium channels, we found that plectasin could dose-dependently block Kv1.3 channel currents through electrophysiological experiments. Besides Kv1.3 channel, plectasin could less inhibit Kv1.1, Kv1.2, IKCa, SKCa3, hERG and KCNQ channels at the concentration of 1 μΜ. Using mutagenesis and channel activation experiments, we found that outer pore region of Kv1.3 channel was the binding site of plectasin, which is similar to the interacting site of Kv1.3 channel recognized by animal toxin blockers. Together, these findings not only highlight the novel function of plectasin as a potassium channel inhibitor, but also imply that defensins from different organisms functionally evolve to be a novel kind of potassium channel inhibitors.

The fungal defensin family enlarged.

Fungi are an emerging source of peptide antibiotics. With the availability of a large number of model fungal genome sequences, we can expect that more and more fungal defensin-like peptides (fDLPs) will be discovered by sequence similarity search. Here, we report a total of 69 new fDLPs encoded by 63 genes, in which a group of fDLPs derived from dermatophytes are defined as a new family (fDEF8) according to sequence and phylogenetic analyses. In the oleaginous fungus Mortierella alpine, fDLPs have undergone extensive gene expansion. Our work further enlarges the fungal defensin family and will help characterize new peptide antibiotics with therapeutic potential.

Fungal defensins, an emerging source of anti-infective drugs

With the rapid increase of antibiotic-resistant pathogens and the decline of discovery and development of new antibiotics, there is an urgent need to exploit alternative anti-infective drugs. Fungal defensin-like peptides are emerging as a class of new source of anti-infective drugs due to their potent antibacterial activity, low toxicity, and high serum stability. Fungal genome sequencing projects combined with the development of recombinant expression techniques will accelerate the discovery of fungal defensin-like peptides.