Lipopeptides from Bacillus velezensis ZLP-101 and their mode of action against bean aphids Acyrthosiphon pisum Harris.

BACKGROUND: Natural products are important sources for the discovery of new biopesticides to control the worldwide destructive pests Acyrthosiphon pisum Harris. Here, insecticidal substances were discovered and characterized from the secondary metabolites of the bio-control microorganism Bacillus velezensis strain ZLP-101, as informed by whole-genome sequencing and analysis. RESULTS: The genome was annotated, revealing the presence of four potentially novel gene clusters and eight known secondary metabolite synthetic gene clusters. Crude extracts, prepared through ammonium sulfate precipitation, were used to evaluate the effects of strain ZLP-101 on Acyrthosiphon pisum Harris aphid pests via exposure experiments. The half lethal concentration (LC50) of the crude extract from strain ZLP-101 against aphids was 411.535 mg/L. Preliminary exploration of the insecticidal mechanism revealed that the crude extract affected aphids to a greater extent through gastric poisoning than through contact. Further, the extracts affected enzymatic activities, causing holes to form in internal organs along with deformation, such that normal physiological activities could not be maintained, eventually leading to death. Isolation and purification of extracellular secondary metabolites were conducted in combination with mass spectrometry analysis to further identify the insecticidal components of the crude extracts. A total of 15 insecticidal active compounds were identified including iturins, fengycins, surfactins, and spergualins. Further insecticidal experimentation revealed that surfactin, iturin, and fengycin all exhibited certain aphidicidal activities, and the three exerted synergistic lethal effects. CONCLUSIONS: This study improved the available genomic resources for B. velezensis and serves as a foundation for comprehensive studies of the insecticidal mechanism by Bacillus velezensis ZLP-101 in addition to the active components within biological control strains.

The Physicochemical and Functional Properties of Biosurfactants: A Review.

Surfactants, also known as surface-active agents, have emerged as an important class of compounds with a wide range of applications. However, the use of chemical-derived surfactants must be restricted due to their potential adverse impact on the ecosystem and the health of human and other living organisms. In the past few years, there has been a growing inclination towards natural-derived alternatives, particularly microbial surfactants, as substitutes for synthetic or chemical-based counterparts. Microbial biosurfactants are abundantly found in bacterial species, predominantly Bacillus spp. and Pseudomonas spp. The chemical structures of biosurfactants involve the complexation of lipids with carbohydrates (glycolipoproteins and glycolipids), peptides (lipopeptides), and phosphates (phospholipids). Lipopeptides, in particular, have been the subject of extensive research due to their versatile properties, including emulsifying, antimicrobial, anticancer, and anti-inflammatory properties. This review provides an update on research progress in the classification of surfactants. Furthermore, it explores various bacterial biosurfactants and their functionalities, along with their advantages over synthetic surfactants. Finally, the potential applications of these biosurfactants in many industries and insights into future research directions are discussed.

Production, characterization, and potential applications of lipopeptides in food systems: A comprehensive review.

Lipopeptides are a class of lipid-peptide-conjugated compounds with differing structural features. This structural diversity is responsible for their diverse range of biological properties, including antimicrobial, antioxidant, and anti-inflammatory activities. Lipopeptides have been attracting the attention of food scientists due to their potential as food additives and preservatives. This review provides a comprehensive overview of lipopeptides, their production, structural characteristics, and functional properties. First, the classes, chemical features, structure-activity relationships, and sources of lipopeptides are summarized. Then, the gene expression and biosynthesis of lipopeptides in microbial cell factories and strategies to optimize lipopeptide production are discussed. In addition, the main methods of purification and characterization of lipopeptides have been described. Finally, some biological activities of the lipopeptides, especially those relevant to food systems along with their mechanism of action, are critically examined.

Exploring the Use of a Lipopeptide in Dipalmitoylphosphatidylcholine Monolayers for Enhanced Detection of Glyphosate in Aqueous Environments.

The growing reliance on pesticides for pest management in agriculture highlights the need for new analytical methods to detect these substances in food and water. Our research introduces a SPRWG-(C18H37) lipopeptide (LP) as a functional analog of acetylcholinesterase (AChE) for glyphosate detection in environmental samples using phosphatidylcholine (PC) monolayers. This LP, containing hydrophilic amino acids linked to an 18-carbon aliphatic chain, alters lipid assembly properties, leading to a more flexible system. Changes included reduced molecular area and peak pressure in Langmuir adsorption isotherms. Small angle X-ray scattering (SAXS) and atomic force microscopy (AFM) analyses provided insights into the LP's structural organization within the membrane and its interaction with glyphosate (PNG). Structural and geometric parameters, as derived from in silico molecular dynamics simulations (MD), substantiated the impact of LP on the monolayer structure and the interaction with PNG. Notably, the presence of the LP and glyphosate increased charge transfer resistance, indicating strong adherence of the monolayer to the indium tin oxide (ITO) surface and effective pesticide interaction. A calibration curve for glyphosate concentration adjustment revealed a detection limit (LOD) of 24 nmol L-1, showcasing the high sensitivity of this electrochemical biosensor. This LOD is significantly lower than that of a similar colorimetric biosensor in aqueous media with a detection limit of approximately 0.3 µmol L-1. Such an improvement in sensitivity likely stems from adding a polar residue to the amino acid chain of the LP.

Production and optimization of surfactin produced from locally isolated Bacillus halotolerans grown on agro-industrial wastes and its antimicrobial efficiency.

INTRODUCTION: Optimal exploitation of the huge amounts of agro-industrial residuals that are produced annually, which endangers the ecosystem and ultimately contributes to climate change, is one of the solutions available to produce value-added compounds. AIM AND OBJECTIVES: This study aimed at the economic production and optimization of surfactin. Therefore, the production was carried out by the microbial conversion of Potato Peel Waste (PPW) and Frying Oil Waste (FOW) utilizing locally isolated Bacillus halotolerans. Also, investigating its potential application as an antimicrobial agent towards some pathogenic strains. RESULTS: Screening the bacterial isolates for surfactin production revealed that the strain with the highest yield (49 g/100 g substrate) and efficient oil displacement activity was genetically identified as B. halotolerans. The production process was then optimized utilizing Central Composite Design (CCD) resulting in the amelioration of yield by 11.4% (from 49 to 55.3 g/100 g substrate) and surface tension (ST) by 8.3% (from 36 to 33 mN/m) with a constant level of the critical micelle concentration (CMC) at 125 mg/L. Moreover, the physiochemical characterization studies of the produced surfactin by FTIR, 1H NMR, and LC-MS/MS proved the existence of a cyclic lipopeptide (surfactin). The investigations further showed a strong emulsification affinity for soybean and motor oil (E24 = 50%), as well as the ability to maintain the emulsion stable over a wide pH (4-10) and temperature (10-100 °C) range. Interestingly, surfactin had a broad-spectrum range of inhibition activity against Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa, klebsiella pneumonia, and Candida albicans. CONCLUSION: Subsequently, the screening of the isolates and the utilized food-processing wastes along with the extraction technique resulted in a high yield of surfactin characterized by acceptable ST and CMC levels. However, optimization of the cultural conditions to improve the activity and productivity was achieved using Factor-At-A-Time (OFAT) and Central Composite Design (CCD). In contrast, surface activity recorded a maximum level of (33 mN/n) and productivity of 55.3 g/100 g substrate. The optimized surfactin had also the ability to maintain the stability of emulsions over a wide range of pH and temperature. Otherwise, the obtained results proved the promising efficiency of the surfactin against bacterial and fungal pathogens.

Nanostructured lipopeptide-based membranomimetics for stabilizing bacteriorhodopsin.

Nanostructured 7-9-residue cyclic and unstructured lipopeptide-based facial detergents have been engineered to stabilize the model integral membrane protein, bacteriorhodopsin. Formation of a cylindrical-type micelle assembly induced by facial amphipathic lipopeptides resembles a biological membrane more effectively than conventional micelles. The hydrophobic face of this cylindrical-type micelle provides extended stability to the membrane protein and the hydrophilic surface interacts with an aqueous environment. In our present study, we have demonstrated experimentally and computationally that lipopeptide-based facial detergents having an unstructured or ß-turn conformation can stabilize membrane proteins. However, constrained peptide detergents can provide enhanced stability to bacteriorhodopsin. In this study, we have computationally examined the structural stability of bacteriorhodopsin in the presence of helical, beta-strand, and cyclic unstructured peptide detergents, and conventional detergent-like peptides. Our study demonstrates that optimal membranomimetics (detergents) for stabilizing a specific membrane protein can be screened based on the following criteria: (i) hydrodynamic radii of the self-assembled peptide detergents, (ii) stability assay of detergent-encased membrane proteins, (iii) percentage covered area of detergent-encased membrane proteins obtained computationally and (iv) protein-detergent interaction energy.

Antibacterial efficacy and membrane mechanism of action of the Serratia-derived non-ionic lipopeptide, serrawettin W2-FL10.

The study aimed to investigate the antibacterial activity, cytotoxicity, and mechanism of action of the non-ionic, cyclic lipopeptide, serrawettin W2-FL10 against Staphylococcus aureus. W2-FL10 exhibited potent activity against the Gram-positive bacteria S. aureus, Enterococcus faecalis, Enterococcus faecium, Listeria monocytogenes, and Bacillus subtilis, with minimum inhibitory concentration (MIC) values ranging from 6.3 to 31.3 µg/mL, while no activity was observed against Gram-negative bacteria. Broth microdilution assays showed that W2-FL10 interacted with key cell membrane components, such as lipid phosphatidyl glycerol and lipoteichoic acid of S. aureus. Upon membrane interaction, W2-FL10 dissipated membrane potential within 12 min and increased S. aureus membrane permeability within 28-40 min, albeit at slower rates and higher concentrations than the lytic peptide melittin. The observed membrane permeability, as detected with propidium iodide (PI), may be attributed to transmembrane pores/lesions, possibly dependent on dimer-driven lipopeptide oligomerization in the membrane. Scanning electron microscopy (SEM) imaging also visually confirmed the formation of lesions in the cell wall of one of the S. aureus strains, and cell damage within 1 h of exposure to W2-FL10, corroborating the rapid time-kill kinetics of the S. aureus strains. This bactericidal action against the S. aureus strains corresponded to membrane permeabilization by W2-FL10, indicating that self-promoted uptake into the cytosol may be part of the mode of action. Finally, this lipopeptide exhibited low to moderate cytotoxicity to the Chinese hamster ovarian (CHO) cell line in comparison to the control (emetine) with an optimal lipophilicity range (log D value of 2.5), signifying its potential as an antibiotic candidate. IMPORTANCE: Antimicrobial resistance is a major public health concern, urgently requiring antibacterial compounds exhibiting low adverse health effects. In this study, a novel antibacterial lipopeptide analog is described, serrawettin W2-FL10 (derived from Serratia marcescens), with potent activity displayed against Staphylococcus aureus. Mechanistic studies revealed that W2-FL10 targets the cell membrane of S. aureus, causing depolarization and permeabilization because of transmembrane lesions/pores, resulting in the leakage of intracellular components, possible cytosolic uptake of W2-FL10, and ultimately cell death. This study provides the first insight into the mode of action of a non-ionic lipopeptide. The low to moderate cytotoxicity of W2-FL10 also highlights its application as a promising therapeutic agent for the treatment of bacterial infections.

Homo and Hetero-Branched Lipopeptide Dendrimers: Synthesis and Antimicrobial Activity.

Di-branched and tetra-branched versions of a previously reported analogue of the lipopeptide battacin were successfully synthesised using thiol-maleimide click and 1, 2, 3-triazole click chemistry. Antimicrobial studies against drug resistant clinical isolates of Escherichia coli (ESBL E. coli Ctx-M14), Pseudomonas aeruginosa (P. aeruginosa Q502), and Methicillin resistant Staphylococcus aureus (MRSA ATCC 33593), as well as clinically isolated Acinetobacter baumannii (A. baumannii ATCC 19606), and P. aeruginosa (ATCC 27853), revealed that the dendrimeric peptides have antimicrobial activity in the low micromolar range (0.5 -- 4 µM) which was 10 times more potent than the monomer peptides. Under high salt concentrations (150 mM NaCl, 2 mM MgCl2, and 2.5 mM CaCl2) the di-branched lipopeptides retained their antimicrobial activity while the monomer peptides were not active (>100 µM). The di-branched triazole click lipopeptide, Peptide 12, was membrane lytic, showed faster killing kinetics, and exhibited antibiofilm activity against A. baumannii and MRSA and eradicated > 85 % preformed biofilms at low micromolar concentrations. The di-branched analogues were > 30-fold potent than the monomers against Candida albicans. Peptide 12 was not haemolytic (HC10 = 932.12 µM) and showed up to 40-fold higher selectivity against bacteria and fungi than the monomer peptide. Peptide 12 exhibited strong proteolytic stability (>80 % not degraded) in rat serum over 24 h whereas > 95 % of the thiol-maleimide analogue (Peptide 10) was degraded. The tetra-branched peptides showed comparable antibacterial potency to the di-branched analogues. These findings indicate that dual branching using triazole click chemistry is a promising strategy to improve the antimicrobial activity and proteolytic stability of battacin based lipopeptides. The information gathered can be used to build effective antimicrobial dendrimeric peptides as new peptide antibiotics.

Relative biomembrane fusogenicities of the tumor-selective liposomes of RGDK- and CGKRK-lipopeptides.

Cancer is the second leading cause of death globally after heart diseases. Currently used highly cytotoxic anti-cancer drugs not only kill cancer cells but also often kill non-cancerous healthy body cells, causing adverse side effects. Efforts are now being directed towards developing tumor-selective chemotherapy. Tumor/tumor endothelial cell selective peptide ligands are being covalently grafted onto the exo-surfaces of drug carriers such as liposomes, polymers, etc. A number of prior studies used conjugation of tumor/tumor endothelial cell-selective RGDK- or CGKRK-peptide ligands on the outer surfaces of liposomes, metal-based nanoparticles, single walled carbon nanotubes (SWNTs), etc. However, studies aimed at examining the relative cell membrane fusogenicities and the relative degrees of cellular uptake for the RGDK- and CGKRK-ligand-grafted nanometric drug carriers have not yet been undertaken. Herein, using the widely used liposomes of DOPC, DOPE, DOPS and cholesterol (45 : 25 : 20 : 15, w/w ratio) as the model biomembranes and the fluorescence resonance energy transfer (FRET) assay for measuring membrane fusogenicities, we show that the liposomes of the RGDK-lipopeptide are more biomembrane fusogenic than the liposomes of the CGKRK-lipopeptide. Notably, such FRET assay-derived relative biomembrane fusogenicities of the liposomes of RGDK- and CGKRK-lipopeptides were found to be consistent with their relative degrees of cellular uptake in cultured cancer cells. The present findings open the door for undertaking in-depth in vivo studies aimed at evaluating the relative therapeutic potential of different nanocarriers of drugs/genes/siRNA having tumor-targeting RGDK- and CGKRK-peptides on their exo-surfaces.

Two promising natural lipopeptides from Bacillus subtilis effectively induced membrane permeabilization in Candida glabrata.

Candida glabrata is an important opportunistic human pathogen well known to develop resistance to antifungal drugs. Due to their numerous desirable qualities, antimicrobial lipopeptides have gained significant attention as promising candidates for antifungal drugs. In the present study, two bioactive lipopeptides (AF4 and AF5 m/z 1071.5 and 1085.5, respectively), coproduced and purified from Bacillus subtilis RLID12.1, consist of seven amino acid residues with lipid moieties. In our previous studies, the reversed phased-HPLC purified lipopeptides demonstrated broad-spectrum of antifungal activities against over 110 Candida albicans, Candida non-albicans and mycelial fungi. Two lipopeptides triggered membrane permeabilization of C. glabrata cells, as confirmed by propidium iodide-based flow cytometry, with PI uptake up to 99% demonstrating fungicidal effects. Metabolic inactivation in treated cells was confirmed by FUN-1-based confocal microscopy. Together, the results indicate that these lipopeptides have potentials to be developed into a new set of antifungals for combating fungal infections.

Deciphering the distinct biocontrol activities of lipopeptides fengycin and surfactin through their differential impact on lipid membranes.

Lipopeptides produced by beneficial bacilli present promising alternatives to chemical pesticides for plant biocontrol purposes. Our research explores the distinct plant biocontrol activities of lipopeptides surfactin (SRF) and fengycin (FGC) by examining their interactions with lipid membranes. Our study shows that FGC exhibits a direct antagonistic activity against Botrytis cinerea and no marked immune-eliciting activity in Arabidopsis thaliana while SRF only demonstrates an ability to stimulate plant immunity. It also reveals that SRF and FGC exhibit diverse effects on membrane integrity and lipid packing. SRF primarily influences membrane physical state without significant membrane permeabilization, while FGC permeabilizes membranes without significantly affecting lipid packing. From our results, we can suggest that the direct antagonistic activity of lipopeptides is linked to their capacity to permeabilize lipid membrane while the stimulation of plant immunity is more likely the result of their ability to alter the mechanical properties of the membrane. Our work also explores how membrane lipid composition modulates the activities of SRF and FGC. Sterols negatively impact both lipopeptides' activities while sphingolipids mitigate the effects on membrane lipid packing but enhance membrane leakage. In conclusion, our findings emphasize the importance of considering both membrane lipid packing and leakage mechanisms in predicting the biological effects of lipopeptides. It also sheds light on the intricate interplay between the membrane composition and the effectiveness of the lipopeptides, providing insights for targeted biocontrol agent design.

Biosurfactant-amphiphilized hyaluronic acid: A dual self-assembly anticancer nanoconjugate and drug vector for synergistic chemotherapy.

Novel amphiphilic nanoconjugates of hyaluronic acid (HA), 50 kDa (HA50) and 100 kDa (HA100), and the lipopeptide biosurfactant surfactin (SF) were developed for potential anticancer applications. Physicochemical characterization indicated the formation of an ester conjugate (HA: SF molar ratio 1: 40) with the HA50-SF derivative exhibiting higher degree of substitution, hydrolytic stability, and surface activity. Self-assembly resulted in nanomicelles with smaller size and greater negative charge relative to SF micelles. Biological data demonstrated distinct anticancer activity of HA50-SF which displayed greater synergistic cytotoxicity and selectivity for MDA-MB 231 and MCF-7 breast cancer cells alongside greater modulation of apoptosis-related biomarkers leading to apoptosis. As bioactive vector for chemotherapeutic agents, the selected HA50-SF nanoconjugate efficiently (70 %) entrapped berberine (BER) producing a sustained release BER-HA50-SF synergistic anticancer nanoformulation. Lactoferrin (Lf) coating for dual HA/Lf targeting endowed Lf/BER-HA50-SF with significantly greater selectivity for both cell lines. A murine Ehrlich breast cancer model provided evidence for the efficacy and safety of Lf/BER-HA50-SF via tumoral, histological, immunohistochemical, molecular and systemic toxicity assessments. Thus, HA-SF nanoconjugates integrating the HA and SF properties and biofunctionalties present a novel biopolymer-biosurfactant platform of benefit to oncology nanomedicine and possibly other applications.

Cinnamosyn, a Cinnamoylated Synthetic-Bioinformatic Natural Product with Cytotoxic Activity.

Most biosynthetic gene clusters (BGCs) are functionally inaccessible by using fermentation methods. Bioinformatic-coupled total synthesis provides an alternative approach for accessing BGC-encoded bioactivities. To date, synthetic bioinformatic natural product (synBNP) methods have focused on lipopeptides containing simple lipids. Here we increase the bioinformatic and synthetic complexity of the synBNP approach by targeting BGCs that encode N-cinnamoyl lipids. This led to our synthesis of cinnamosyn, a 10-mer N-cinnamoyl-containing peptide that is cytotoxic to human cells.

Bacillus velezensis iturins inhibit the hemolytic activity of Staphylococcus aureus.

Bovine mastitis caused by S. aureus has a major economic impact on the dairy sector. With the crucial need for new therapies, anti-virulence strategies have gained attention as alternatives to antibiotics. Here we aimed to identify novel compounds that inhibit the production/activity of hemolysins, a virulence factor of S. aureus associated with mastitis severity. We screened Bacillus strains obtained from diverse sources for compounds showing anti-hemolytic activity. Our results demonstrate that lipopeptides produced by Bacillus spp. completely prevented the hemolytic activity of S. aureus at certain concentrations. Following purification, both iturins, fengycins, and surfactins were able to reduce hemolysis caused by S. aureus, with iturins showing the highest anti-hemolytic activity (up to 76% reduction). The lipopeptides showed an effect at the post-translational level. Molecular docking simulations demonstrated that these compounds can bind to hemolysin, possibly interfering with enzyme action. Lastly, molecular dynamics analysis indicated general stability of important residues for hemolysin activity as well as the presence of hydrogen bonds between iturins and these residues, with longevous interactions. Our data reveals, for the first time, an anti-hemolytic activity of lipopeptides and highlights the potential application of iturins as an anti-virulence therapy to control bovine mastitis caused by S. aureus.

Noducyclamides A1-A4, B1, and B2 from the Cyanobacterium Nodularia sp. NIES-3585.

A chemical investigation of the hydrophilic fraction of a cultured Nodularia sp. (NIES-3585) afforded six new cyclic lipopeptides, noducyclamides A1-A4 (1-4) containing 10 amino acid residues and dodecapeptides noducyclamides B1 and B2 (5 and 6). The planar structures of these lipopeptides were elucidated based on the combination of HRMS and 1D and 2D NMR spectroscopic data analyses. These peptides are structurally analogous to laxaphycins and contain the nonproteinogenic amino acids 3-hydroxyvaline and 3-hydroxyleucine and a ß-amino decanoic acid residue. The absolute configurations of the noducyclamides (1-6) were determined by acid hydrolysis, followed by advanced Marfey's analysis. Noducyclamide B1 (5) showed cytotoxic activities against MCF7 breast cancer cell lines with an IC50 value of 3.0 µg/mL (2.2 µM).

Multifunctional Lipidated Protein Carrier with a Built-In Adjuvant as a Universal Vaccine Platform Potently Elevates Immunogenicity of Weak Antigens.

Weak antigens represented by MUC1 are poorly immunogenic, which greatly constrains the development of relevant vaccines. Herein, we developed a multifunctional lipidated protein as a carrier, in which the TLR1/2 agonist Pam3CSK4 was conjugated to the N-terminus of MUC1-loaded carrier protein BSA through pyridoxal 5'-phosphate-mediated transamination reaction. The resulting Pam3CSK4-BSA-MUC1 conjugate was subsequently incorporated into liposomes, which biomimics the membrane structure of tumor cells. The results indicated that this lipidated protein carrier significantly enhanced antigen uptake by APCs and obviously augmented the retention of the vaccine at the injection site. Compared with the BSA-MUC1 and BSA-MUC1 + Pam3CSK4 groups, Pam3CSK4-BSA-MUC1 evoked 22- and 11-fold increases in MUC1-specific IgG titers. Importantly, Pam3CSK4-BSA-MUC1 elicited robust cellular immunity and significantly inhibited tumor growth. This is the first time that lipidated protein was constructed to enhance antigen immunogenicity, and this universal carrier platform exhibits promise for utilization in various vaccines, holding the potential for further clinical application.

Discovery and Derivatization of Tridecaptin Antibiotics with Altered Host Specificity and Enhanced Bioactivity.

The prevalence of multidrug-resistant (MDR) pathogens combined with a decline in antibiotic discovery presents a major challenge for health care. To refill the discovery pipeline, we need to find new ways to uncover new chemical entities. Here, we report the global genome mining-guided discovery of new lipopeptide antibiotics tridecaptin A5 and tridecaptin D, which exhibit unusual bioactivities within their class. The change in the antibacterial spectrum of Oct-TriA5 was explained solely by a Phe to Trp substitution as compared to Oct-TriA1, while Oct-TriD contained 6 substitutions. Metabolomic analysis of producer Paenibacillus sp. JJ-21 validated the predicted amino acid sequence of tridecaptin A5. Screening of tridecaptin analogues substituted at position 9 identified Oct-His9 as a potent congener with exceptional efficacy against Pseudomonas aeruginosa and reduced hemolytic and cytotoxic properties. Our work highlights the promise of tridecaptin analogues to combat MDR pathogens.

Production of Lipopeptides from Bacillus velezensis BVQ121 and Their Application in Chitosan Antibacterial Coating.

The extracellular substance of Bacillus has antibacterial effects inhibiting multiple foodborne pathogens and plays important roles in food production. This study found one Bacillus velezensis BVQ121 strain producing antibacterial lipopeptides (BVAL). After optimization of the fermentation conditions, the BVAL yield was the highest at 1.316 ± 0.03 g/L in reality with the initial pH 6.0, temperature 31 °C, and shaker speed 238 rpm when the optimal nitrogen and carbon sources were used in Landy medium for fermentation. The antibacterial components were identified as iturin, surfactin, and fengycin by HPLC and MALDI-TOF-MS. The MIC was at 2 mg/mL and MBC was at 5 mg/mL. The 6% weight ratio of nanocellulose dosage in chitosan solution could improve the tensile length and strength of the film, and the antibacterial performance was enhanced by the addition of BVAL. The addition of BVAL had no effect on the color and ductility of the film and improved its antibacterial effect. The shelf life of pigeon eggs can be extended by more than 10 days to resist bacterial infections after coating with the chitosan-nanocellulose-BVAL film solution.

Cholesterol as an inbuilt immunoadjuvant for a lipopeptide vaccine against group A Streptococcus infection.

Peptide-based vaccines can trigger highly specific immune responses, although peptides alone are usually unable to confer strong humoral or cellular immunity. Consequently, peptide antigens are administered with immunostimulatory adjuvants, but only a few are safe and effective for human use. To overcome this obstacle, herein a peptide antigen was lipidated to effectively anchor it to liposomes and emulsion. A peptide antigen B cell epitope from Group A Streptococcus M protein was conjugated to a universal T helper epitope, the pan DR-biding epitope (PADRE), alongside a lipidic moiety cholesterol. Compared to a free peptide antigen, the lipidated version (LP1) adopted a helical conformation and self-assembled into small nanoparticles. Surprisingly, LP1 alone induced the same or higher antibody titers than liposomes or emulsion-based formulations. In addition, antibodies produced by mice immunized with LP1 were more opsonic than those induced by administering the antigen with incomplete Freund's adjuvant. No side effects were observed in the immunized mice and no excessive inflammatory immune responses were detected. Overall, this study demonstrated how simple conjugation of cholesterol to a peptide antigen can produce a safe and efficacious vaccine against Group A Streptococcus - the leading cause of superficial infections and the bacteria responsible for deadly post-infection autoimmune disorders.

Total synthesis of antifungal lipopeptide iturin A analogues and evaluation of their bioactivity against F. graminearum.

The pursuit of novel antifungal agents is imperative to tackle the threat of antifungal resistance, which poses major risks to both human health and to food security. Iturin A is a cyclic lipopeptide, produced by Bacillus sp., with pronounced antifungal properties against several pathogens. Its challenging synthesis, mainly due to the laborious synthesis of the ß-amino fatty acid present in its structure, has hindered the study of its mode of action and the development of more potent analogues. In this work, a facile synthesis of bioactive iturin A analogues containing an alkylated cysteine residue is presented. Two analogues with opposite configurations of the alkylated cysteine residue were synthesized, to evaluate the role of the stereochemistry of the newly introduced amino acid on the bioactivity. Antifungal assays, conducted against F. graminearum, showed that the novel analogues are bioactive and can be used as a synthetic model for the design of new analogues and in structure-activity relationship studies. The assays also highlight the importance of the ß-amino acid in the natural structure and the role of the stereochemistry of the amino fatty acid, as the analogue with the D configuration showed stronger antifungal properties than the one with the L configuration.