Natural Antimicrobial Peptides Research Articles

Innovative perspectives on bacteriocins: advances in classification, synthesis, mode of action, and food industry applications.

Bacteriocins, natural antimicrobial peptides produced by bacteria, present eco-friendly, non-toxic, and cost-effective alternatives to traditional chemical antimicrobial agents in the food industry. This review provides a comprehensive update on the classification of bacteriocins in food preservation. It highlights the significant industrial potential of pediocin-like and two-peptide bacteriocins, emphasizing chemical synthesis methods like Fmoc-SPPS to meet the demand for bioactive bacteriocins. The review details the mode of action, focusing on mechanisms such as transmembrane potential disruption and pH-dependent effects. Furthermore, it addresses the limitations of bacteriocins in food preservation and explores the potential of nanotechnology-based encapsulation to enhance their antimicrobial efficacy. The benefits of nanoencapsulation, including improved stability, extended antimicrobial spectrum, and enhanced functionality, are underscored. This understanding is crucial for advancing the application of bacteriocins to ensure food safety and quality.

Efflux Pump Inhibitors as a Broad-Spectrum Strategy to Combat Multidrug Resistance Acenitobacter Baumanii

The World Health Organization (WHO) identifies A. baumannii as one of the most resistant pathogens. Over the past few years, there has been a significant rise in its resistance to a wide array of antibiotics. Multidrug resistance is attributed to several mechanisms, with efflux pumps being the most notable. Recently, the development of new treatment strategies to tackle A. baumannii infections has shown great promise. This review focuses on the various families of efflux pumps in A. baumannii and explores innovative therapeutic options that have emerged over the past four years. Specifically, we emphasize synthetic and natural efflux pump inhibitors (EPIs), nanoparticles, bacteriophage therapy, and antimicrobial peptides (AMPs) as part of these novel strategies.

Application of self-assembled antibacterial nanofiber loaded oriented artificial skin in infected diabetic-related wound regeneration

Diabetic patients develop wounds that exhibit delayed healing, prolonged inflammatory responses, and slower epithelialization kinetics compared to non-diabetic patients. Diabetic foot ulcers(DFUs) affect approximately 18.6 million people worldwide. The presence of a high glucose microenvironment in DFUs results in the significant accumulation of bacterial infection and advanced glycation end products (AGEs). To solve this, a self-assemble antibacterial nanofiber(ANF) loaded oriential artificial skin (ANF@OAS) was introduced in this research, which is consisted of L/D-phenylalanine derivatives coupled the natural antimicrobial peptides. The ANF@OAS can effectively reduce AGEs production and suppress multiple resistant bacteria. Additionally, the ANF@OAS can suppress infection and stimulate wound healing in infected diabetic mice.

Durian rind pectin blended with nisin coating to preserve egg quality and reduce bacteria

In the absence of refrigerated storage, coating materials may offer effective alternatives for preserving egg shelf life. Durian rind, rich in pectin, seals eggshell pores, while nisin, a natural antimicrobial peptide, helps reduce bacteria. This study investigated the effectiveness of durian rind pectin combined with nisin to preserve egg quality and reduce bacterial contamination. Eggs were divided by treatment: (1) uncoated or control; (2) durian rind pectin coated; and (3) coated with durian rind pectin and nisin. All eggs were kept at room temperature (25–30 °C) for five weeks and evaluated weekly for weight loss, Haugh unit (HU), yolk index (YI), egg white pH, and total bacterial count on eggshells and egg content. Results were that durian rind pectin coating, with or without nisin, significantly reduced weight loss (8.5–8.7 % in coated eggs versus 10.5 % in uncoated eggs by day 35); maintained Grade A HU until day 28 (uncoated eggs dropped to Grade B by day 14); and preserved a higher YI with a lower egg white pH compared to uncoated eggs. The total bacterial count on eggshells was lowest in eggs coated with durian rind pectin and nisin (4.06 log10 CFU/mL), compared to durian rind pectin alone (5.15 log10 CFU/mL) and uncoated eggs (5.85 log10 CFU/mL) at day 35. These findings suggest that durian rind pectin coating may extend egg shelf life up to 28 days by maintaining a Grade A HU and an egg white pH under 9.2. Adding nisin enhances antibacterial activity, reducing bacterial count on eggshells.

Human Cathelicidin, LL-37 a potential antiviral therapeutic for Rift Valley Fever Virus in Egypt.

Rift Valley Fever Virus (RVFV) is an arbovirus that circulates among animals and can be transmitted to humans. Mosquitoes are the primary vectors that allow RVFV to spread vertically and horizontally. Egypt was exposed to frequent outbreaks with devastating economic consequences. RVFV has a high incidence of worldwide dissemination and no specific vaccine or therapy. Linear Human Cathelicidin (LL-37), is a natural antimicrobial peptide with antiviral activity against numerous viruses. In addition to immunomodulatory effects, LL-37 directly influences viral encapsulation. This study aimed to evaluate the antiviral activity of LL-37 against RVFV in vitro. The post-entry and pre-incubation of LL-37 within Vero cells were assessed in the absence and presence of RVFV. LL-37 activity was assessed using a TCID50 endpoint test, qRT-PCR, and a western blot. When genomic RVFV was quantified, it resulted in a 48% direct inactivation of the viral envelope and a 36% reduction when the virus was pre-incubated with LL-37 before infection. LL-37 decreased viral infection by 75% and protected Vero cells against RVFV infection by 47% at a 1.25 µg/ml dosage. These findings imply that LL-37 exerts antiviral efficacy against RVFV by restricting virus entrance through direct disruption of the virus envelope and indirectly by triggering an immunological response. The effect of LL-37 is time-dependent. As a result, LL-37 may provide rapid and affordable therapies for RVFV infection in Egypt, both during outbreaks and as a preventive strategy.

Self-Assembling and Pore-Forming Peptoids as Antimicrobial Biomaterials.

Bacterial infections have been a serious threat to mankind throughout history. Natural antimicrobial peptides (AMPs) and their membrane disruption mechanism have generated immense interest in the design and development of synthetic mimetics that could overcome the intrinsic drawbacks of AMPs, such as their susceptibility to proteolytic degradation and low bioavailability. Herein, by exploiting the self-assembly and pore-forming capabilities of sequence-defined peptoids, we discovered a family of low-molecular weight peptoid antibiotics that exhibit excellent broad-spectrum activity and high selectivity toward a panel of clinically significant Gram-positive and Gram-negative bacterial strains, including vancomycin-resistant Enterococcus faecalis (VREF), methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant Staphylococcus epidermidis (MRSE), Escherichia coli, Pseudomonas aeruginosa, and Klebsiella pneumoniae. Tuning the peptoid side chain chemistry and structure enabled us to tune the efficacy of antimicrobial activity. Mechanistic studies using transmission electron microscopy (TEM), bacterial membrane depolarization and lysis, and time-kill kinetics assays along with molecular dynamics simulations reveal that these peptoids kill both Gram-positive and Gram-negative bacteria through a membrane disruption mechanism. These robust and biocompatible peptoid-based antibiotics can provide a valuable tool for combating emerging drug resistance.

Unlocking the Potential: PEGylation and Molecular Weight Reduction of Ionenes for Enhanced Antifungal Activity and Biocompatibility.

Numerous synthetic polymers, imitating natural antimicrobial peptides, have demonstrated potent antimicrobial activity, positioning them as potential candidates for new antimicrobial drugs. However, the high activity of these molecules often comes at the cost of elevated toxicity against eukaryotic organisms. In this study, a series of cationic ionenes with varying molecular weights to assess the influence of polymer chain length on ionene activity is investigated. To enhance polymer antimicrobial activity and limit toxicity a PEG side chain is introduced into the repeating unit. The resulting molecules consistently exhibited high activity against three model organisms: E. coli, S. aureus and C. albicans. The incorporation of side PEG chain improves antifungal properties and biocompatibility, regardless of molecular weight. The most important finding of this work is that the reduction of polymer molecular mass led to increased antifungal activity and reduced cytotoxicity against HMF and MRC-5 cell lines simultaneously. As a result, the best-performing molecules reported herein displayed minimal inhibitory concentrations (MIC) as low as 2 and 0.0625µgmL1 for C. albicans and C. tropicalis respectively, demonstrating exceptional selectivity. It is plausible that some of described herein molecules can serve as potential lead candidates for new antifungal drugs.

Stable peptide-assembled nanozyme mimicking dual antifungal actions

Natural antimicrobial peptides (AMPs) and enzymes (AMEs) are promising non-antibiotic candidates against antimicrobial resistance but suffer from low efficiency and poor stability. Here, we develop peptide nanozymes which mimic the mode of action of AMPs and AMEs through de novo design and peptide assembly. Through modelling a minimal building block of IHIHICI is proposed by combining critical amino acids in AMPs and AMEs and hydrophobic isoleucine to conduct assembly. Experimental validations reveal that IHIHICI assemble into helical β-sheet nanotubes with acetate modulation and perform phospholipase C-like and peroxidase-like activities with Ni coordination, demonstrating high thermostability and resistance to enzymatic degradation. The assembled nanotubes demonstrate cascade antifungal actions including outer mannan docking, wall disruption, lipid peroxidation and subsequent ferroptotic death, synergistically killing >90% Candida albicans within 10 min on disinfection pad. These findings demonstrate an effective de novo design strategy for developing materials with multi-antimicrobial mode of actions.

Prevention of premature birth in female patients with bacterial vaginosis using a complex of natural antimicrobial peptides and cytokines

Introduction. The prevalence of bacterial vaginosis (BV) comprises 23–29 %, which in pregnant women is a known risk factor for premature birth (PB) that rates increases by 2.9-fold. BV treatment with antibiotics has no effect PB incidence, therefore stressing a need to search for alternative remedies.Aim: to evaluate the effectiveness of treatment, including antibacterial therapy and a complex preparation containing natural antimicrobial peptides and cytokines, to reduce the incidence of birth defects in pregnant women with BV.Materials and Methods. Design: a prospective open comparative cohort study in parallel groups was conducted with 101 pregnant women: Group I (n = 69) received the antibiotic Metronidazole, 500 mg tablets orally twice a day for 7 days, and a complex preparation containing exogenous natural antimicrobial peptides and cytokines (Superlymph®) suppositories per 25 IU once a day vaginally in the evening for 20 days; Group II (n = 32) received Metronidazole alone (the same regimen). Patient examination was carried out using approaches included clinical methods, accepted in obstetrics, and laboratory tests – microscopy of vaginal content smears, real-time polymerase chain reaction (PCR).Results. The PB (within 240–366 weeks) incidence in Group I was significantly lower than in Group II and comprised 2.9 % vs. 21.9 %, respectively (p = 0.004), with an 8-fold decline in developing PB risk (relative risk (RR) = 0.13; 95 % confidence interval (CI) = 0.03–0.60), whereas inter-group percentage of pregnant women with high risk (PB history) was comparable (p = 0.39) so that PB incidence did not differ from pregnant women without former PB. Microbiological recovery after treatment for BV was achieved in 85.5 % of patients from Group I vs. 56.3 % in Group II (RR = 1.52; 95 % CI = 1.10–2.10; p = 0.002) based on real-time PCR data. The persistence of anaerobic flora after treatment was significantly lower in Group I vs. Group II reaching 7.2 and 34.4 % (p < 0.001), respectively, with a 5-fold lower PB risk (RR = 0.21; 95 % CI = 0.08–0.56). Cessation of viral shedding compared to the number of patients with initial viral shedding was achieved in 94.6 % vs. 8.3 % of patients, respectively, with a 50-fold decline in risk (RR = 0.02; 95 % CI = 0.005–0.08; p < 0.001). The number of newborns weighing less than 2500 g was significantly lower from paired mothers who received Superlymph® + Metronidazole comprising 2.9 % vs. 15.6 % treated with Metronidazole alone (p = 0.03), whereas a risk of low birth weight neonates was decreased by 6-fold (RR = 0.16; 95 % CI = 0.03–0.88). The condition of the neonates assessed by birth Apgar score was comparable.Conclusion. The use of a complex preparation Superlymph® (suppositories per 25 IU once an day vaginally, for 20 days) along with oral antibiotic Мetronidazole in pregnant women with BV facilitates a decline in PB incidence down to 2.9 % at gestational age of 240–366 weeks lowering a risk of PB exceeding that of antibacterial therapy by 8-fold, including patients with former PB.

A Cyclic Peptide Based on Pheasant Cathelicidin Inhibits Influenza A H1N1 Virus Infection.

Influenza viruses are the leading cause of upper respiratory tract infections, leading to several global pandemics and threats to public health. Due to the continuous mutation of influenza A viruses, there is a constant need for the development of novel antiviral therapeutics. Recently, natural antimicrobial peptides have provided an opportunity for the discovery of anti-influenza molecules. Here, we designed several peptides based on pheasant cathelicidin and tested their antiviral activities and mechanisms against the H1N1 virus. Of note, the designed peptides Pc-4 and Pc-5 were found to inhibit replication of the H1N1 virus with an IC50 = 8.14 ± 3.94 µM and 2.47 ± 1.95 µM, respectively. In addition, the cyclic peptide Pc-5 was found to induce type I interferons and the expression of interferon-induced genes. An animal study showed that the cyclic peptide Pc-5 effectively inhibited H1N1 virus infection in a mouse model. Taken together, our work reveals a strategy for designing cyclic peptides and provides novel molecules with therapeutic potential against influenza A virus infection.

Exploring the Efficacy of Peptides and Mimics against Influenza A Virus, Adenovirus, and Murine Norovirus.

The ongoing battle against viral pandemics continues, with the possibility of future outbreaks. The search for effective antiviral compounds that can combat a diverse range of viruses continues to be a focal point of research. This study investigated the efficacy of two natural antimicrobial peptides (AMPs) (lactoferricin and LL-37), two synthetic AMPs (melimine and Mel4), and nine AMP mimics (758, 1091, 1096, 1083, 610, NAPL, 3-BIPL, 4-BIPL, and Sau-22) against influenza A virus strains H1N1 and H3N2, human adenovirus 5 (HAdV-5), and murine norovirus 1 (MNV-1). These compounds were tested using virus pre-treatment, cell pre-treatment, or post-cell entry treatment assays, electron microscopy, and circular dichroism (CD), alongside evaluations of cytotoxicity against the host cells. After virus pre-treatment, the AMP mimics 610 and Sau-22 had relatively low IC50 values for influenza strains H1N1 (2.35 and 6.93 µM, respectively) and H3N2 (3.7 and 5.34 µM, respectively). Conversely, natural and synthetic AMPs were not active against these strains. For the non-enveloped viruses, the AMP Mel4 and mimic 1083 had moderate activity against HAdV-5 (Mel4 IC50 = 47.4 µM; 1083 IC50 = 47.2 µM), whereas all AMPs, but none of the mimics, were active against norovirus (LL-37 IC50 = 4.2 µM; lactoferricin IC50 = 23.18 µM; melimine IC50 = 4.8 µM; Mel4 IC50 = 8.6 µM). Transmission electron microscopy demonstrated that the mimics targeted the outer envelope of influenza viruses, while the AMPs targeted the capsid of non-enveloped viruses. CD showed that Mel4 adopted an α-helical structure in a membrane mimetic environment, but mimic 758 remained unstructured. The diverse activity against different virus groups is probably influenced by charge, hydrophobicity, size, and, in the case of natural and synthetic AMPs, their secondary structure. These findings underscore the potential of peptides and mimics as promising candidates for antiviral therapeutics against both enveloped and non-enveloped viruses.

Recent advances in the therapeutic potential of cathelicidins.

The alarming increase in antimicrobial resistance in the last decades has prompted the search for alternatives to control infectious diseases. Antimicrobial peptides (AMPs) represent a heterogeneous class of molecules with ample antibacterial, antiviral, and antifungal effects. They can be found in many organisms, including all classes of vertebrates, providing a valuable source of new antimicrobial agents. The unique properties of AMPs make it harder for microbes develop resistance, while their immunomodulatory properties and target diversity reinforce their translational use in multiple diseases, from autoimmune disorders to different types of cancer. The latest years have witnessed a vast number of studies evaluating the use of AMPs in therapy, with many progressing to clinical trials. The present review explores the recent developments in the medicinal properties of cathelicidins, a vast family of AMPs with potent antimicrobial and immunomodulatory effects. Cathelicidins from several organisms have been tested in disease models of viral and bacterial infections, inflammatory diseases, and tumors, with encouraging results. Combining nanomaterials with active, natural antimicrobial peptides, including LL-37 and synthetic analogs like ceragenins, leads to the creation of innovative nanoagents with significant clinical promise. However, there are still important limitations, such as the toxicity of many cathelicidins to healthy host cells and low stability in vivo. The recent advances in nanomaterials and synthetic biology may help overcome the current limitations, enabling the use of cathelicidins in future therapeutics. Furthermore, a better understanding of the mechanisms of cathelicidin action in vivo and their synergy with other host molecules will contribute to the development of safer, highly effective therapies.

Antifungal Plant Defensins as an Alternative Tool to Combat Candidiasis.

Currently, the spread of fungal infections is becoming an urgent problem. Fungi of the Candida genus are opportunistic microorganisms that cause superficial and life-threatening systemic candidiasis in immunocompromised patients. The list of antifungal drugs for the treatment of candidiasis is very limited, while the prevalence of resistant strains is growing rapidly. Therefore, the search for new antimycotics, including those exhibiting immunomodulatory properties, is of great importance. Plenty of natural compounds with antifungal activities may be extremely useful in solving this problem. This review evaluates the features of natural antimicrobial peptides, namely plant defensins as possible prototypes of new anticandidal agents. Plant defensins are important components of the innate immune system, which provides the first line of defense against pathogens. The introduction presents a brief summary regarding pathogenic Candida species, the pathogenesis of candidiasis, and the mechanisms of antimycotic resistance. Then, the structural features of plant defensins, their anticandidal activities, their mechanisms of action on yeast-like fungi, their ability to prevent adhesion and biofilm formation, and their combined action with conventional antimycotics are described. The possible mechanisms of fungal resistance to plant defensins, their cytotoxic activity, and their effectiveness in in vivo experiments are also discussed. In addition, for the first time for plant defensins, knowledge about their immunomodulatory effects is also presented.

Antimicrobial activity and mechanism of chitosan-coated epsilon-poly-lysine nanoliposomes against Alicyclobacillus spp. in apple juice

Epsilon-poly-lysine (EPL), a natural cationic antimicrobial peptide, is widely used in food processing and preservation. However, when applied to fruit and vegetable juice, EPL can cause turbidity and precipitation, which reduces the antimicrobial properties and juice quality. As a colloidal delivery system, nanoliposome provides a novel insight into improve the activity of EPL. In this study, the antibacterial activity and mechanism of chitosan-coated EPL nanoliposomes (CS-EPLLP) against Alicyclobacillus spp. in apple juice were evaluated. It was found that the minimum inhibitory concentration of CS-EPLLP against the strains of A. acidocaldarius, A. fructus and A. acidoterrestris were in the ranges of 0.625–1.25 mg/L, while the minimum bactericidal concentration of CS-EPLLP against these strains in apple juice were 0.625–2.5 mg/L. Compared to A. fructus and A. acidoterrestris, A. acidocaldarius displayed a better resistance to CS-EPLLP. Meanwhile, the modification of chitosan can improve the bactericidal effect of EPL and EPL nanoliposomes. In addition, CS-EPLLP treatment can lead to the inactivation of bacteria cells by causing surface damage, destroying their membrane integrity, preventing the intracellular ATP synthesis, increasing intracellular protein leakage and cells membrane permeability. Furthermore, the addition of CS-EPLLP in the amounts of MBC had no significant effect on the color change, transmittance, centrifuge sedimentation rate and main volatile compounds of apple juice. The prepared CS-EPLLP and established method provides a new insight into the inactivation of bacteria in fruit juice.

Lysine-homologue substitution: Impact on antimicrobial activity and proteolytic stability of cationic stapled heptapeptides

Numerous natural antimicrobial peptides (AMPs) exhibit a cationic amphipathic helical conformation, wherein cationic amino acids, such as lysine and arginine, play pivotal roles in antimicrobial activity by aiding initial attraction to negatively charged bacterial membranes. Expanding on our previous work, which introduced a de novo design of amphipathic helices within cationic heptapeptides using an 'all-hydrocarbon peptide stapling' approach, we investigated the impact of lysine-homologue substitution on helix formation, antimicrobial activity, hemolytic activity, and proteolytic stability of these novel AMPs. Our results demonstrate that substituting lysine with ornithine enhances both the antimicrobial activity and proteolytic stability of the stapled heptapeptide AMP series, while maintaining low hemolytic activity. This finding underscores lysine-homologue substitution as a valuable strategy for optimizing the therapeutic potential of diverse cationic AMPs.

Unveiling novel anti-viral mechanisms of ε-poly-l-lysine on tobacco mosaic virus-infected Nicotiana tabacum through microRNA and transcriptome sequencing

MicroRNAs (miRNAs) play important roles in plant defense against various pathogens. ε-poly-l-lysine (ε-PL), a natural anti-microbial peptide produced by microorganisms, effectively suppresses tobacco mosaic virus (TMV) infection. To investigate the anti-viral mechanism of ε-PL, the expression profiles of miRNAs in TMV-infected Nicotiana tabacum after ε-PL treatment were analyzed. The results showed that the expression levels of 328 miRNAs were significantly altered by ε-PL. Degradome sequencing was used to identify their target genes. Integrative analysis of miRNAs target genes and gene-enriched GO/KEGG pathways indicated that ε-PL regulates the expression of miRNAs involved in critical pathways of plant hormone signal transduction, host defense response, and plant pathogen interaction. Subsequently, virus induced gene silencing combined with the short tandem targets mimic technology was used to analyze the function of these miRNAs and their target genes. The results indicated that silencing miR319 and miR164 reduced TMV accumulation in N. benthamiana, indicating the essential roles of these miRNAs and their target genes during ε-PL-mediated anti-viral responses. Collectively, this study reveals that microbial source metabolites can inhibit plant viruses by regulating crucial host miRNAs and further elucidate anti-viral mechanisms of ε-PL.

Antiviral Effect of Antimicrobial Peptoid TM9 and Murine Model of Respiratory Coronavirus Infection.

New antiviral agents are essential to improving treatment and control of SARS-CoV-2 infections that can lead to the disease COVID-19. Antimicrobial peptoids are sequence-specific oligo-N-substituted glycine peptidomimetics that emulate the structure and function of natural antimicrobial peptides but are resistant to proteases. We demonstrate antiviral activity of a new peptoid (TM9) against the coronavirus, murine hepatitis virus (MHV), as a closely related model for the structure and antiviral susceptibility profile of SARS-CoV-2. This peptoid mimics the human cathelicidin LL-37, which has also been shown to have antimicrobial and antiviral activity. In this study, TM9 was effective against three murine coronavirus strains, demonstrating that the therapeutic window is large enough to allow the use of TM9 for treatment. All three isolates of MHV generated infection in mice after 15 min of exposure by aerosol using the Madison aerosol chamber, and all three viral strains could be isolated from the lungs throughout the 5-day observation period post-infection, with the peak titers on day 2. MHV-A59 and MHV-A59-GFP were also isolated from the liver, heart, spleen, olfactory bulbs, and brain. These data demonstrate that MHV serves as a valuable natural murine model of coronavirus pathogenesis in multiple organs, including the brain.

High-yield and cost-effective biosynthesis process for producing antimicrobial peptide AA139

AA139, a variant of natural antimicrobial peptide (AMP) arenicin-3, displayed potent activity against multidrug-resistant (MDR) and extensively drug-resistant (XDR) Gram-negative bacteria. Nevertheless, there were currently few reports on the bioprocess of AA139, and the yields were less than 5 mg/L. Additionally, it was difficult and expensive to prepare AA139 through chemical synthesis due to its complex structure. These factors have impeded the further research and following clinical application of AA139. Here, we reported a bioprocess for the preparation of AA139, which was expressed in Escherichia coli (E. coli) BL21 (DE3) intracellularly in a soluble form via SUMO (small ubiquitin-related modifier) fusion technology. Then, recombinant AA139 (rAA139, refer to AA139 obtained by recombinant expression in this study) was obtained through the simplified downstream process, which was rationally designed in accordance with the physicochemical characteristics. Subsequently, the expression level of the interest protein was increased by 54% after optimization of high cell density fermentation (HCDF). Finally, we obtained a yield of 56 mg of rAA139 from 1 L culture with a purity of 98%, which represented the highest reported yield of AA139 to date. Furthermore, various characterizations were conducted to confirm the molecular mass, disulfide bonds, and antimicrobial activity of rAA139.

Cathelicidin boosts the antifungal activity of neutrophils and improves prognosis during Aspergillus fumigatus keratitis.

Aspergillus fumigatus (A. fumigatus) is one of the common pathogens of fungal keratitis. Fungal growth and invasion cause excessive inflammation and corneal damage, leading to severe vision loss. Neutrophils are the primary infiltrating cells critical for fungal clearance. Cathelicidin [LL-37 in humans and cathelicidin-related antimicrobial peptide (CRAMP) in mice], a natural antimicrobial peptide, can directly inhibit the growth of many pathogens and regulate immune responses. However, the role of cathelicidin and its effect on neutrophils in A. fumigatus keratitis remain unclear. By establishing A. fumigatus keratitis mouse models, we found that cathelicidin was increased in A. fumigatus keratitis. It could reduce fungal loads, lower clinical scores, and improve corneal transparency. Restriction of CRAMP on fungal proliferation was largely counteracted in CD18-/- mice, in which neutrophils cannot migrate into infected sites. When WT neutrophils were transferred into CD18-/- mice, corneal fungal loads were distinctly reduced, indicating that neutrophils are vital for CRAMP-mediated resistance. Furthermore, cathelicidin promoted neutrophils to phagocytose and degrade conidia both in vitro and in vivo. CXC chemokine receptor 2 (CXCR2) was reported to be a functional receptor of LL-37 on neutrophils. CXCR2 antagonist SB225002 or phospholipase C (PLC) inhibitor U73122 weakened LL-37-induced phagocytosis. Meanwhile, LL-37 induced PLC γ phosphorylation, which was attenuated by SB225002. SB225002 or the autophagy inhibitors Bafilomycin-A1 and 3-Methyladenine weakened LL-37-induced degradation of conidia. Transmission electron microscopy (TEM) observed that LL-37 increased autophagosomes in Aspergillus-infected neutrophils. Consistently, LL-37 elevated autophagy-associated protein expressions (Beclin-1 and LC3-II), but this effect was weakened by SB225002. Collectively, cathelicidin reduces fungal loads and improves the prognosis of A. fumigatus keratitis. Both in vitro and in vivo, cathelicidin promotes neutrophils to phagocytose and degrade conidia. LL-37/CXCR2 activates PLC γ to amplify neutrophils' phagocytosis and induces autophagy to eliminate intracellular conidia.

Anti-Staphylococcus aureus effects of natural antimicrobial peptides and the underlying mechanisms.

Staphylococcus aureus can cause localized infections such as abscesses and pneumonia, as well as systemic infections such as bacteremia and sepsis. Especially, methicillin-resistant S. aureus often presents multidrug resistance, which becomes a major clinical challenge. One of the most common reasons for methicillin-resistant S. aureus antibiotic resistance is the presence of biofilms. Natural antimicrobial peptides derived from different species have shown effectiveness in combating S. aureus biofilms. In this review, we summarize the inhibitory activity of antimicrobial peptides against S. aureus planktonic cells and biofilms. We also summarize the possible inhibitory mechanisms, involving cell adhesion inhibition, membrane fracture, biofilm disruption and DNA disruption. We believe this can provide the basis for further research against S. aureus biofilm-associated infections.