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Equine endometritis is one of the main causes of subfertility in the mare. Unraveling the molecular mechanisms involved in this condition and pinpointing proteins with biomarker potential could be crucial in both diagnosing and treating this condition. This study aimed to identify the endometritis-induced changes in the endometrial proteome in mares and to elucidate potential biological processes in which these proteins may be involved. Secondly, biomarkers related to bacterial endometritis (BE) in mares were identified. Uterine lavage fluid samples were collected from 28 mares (14 healthy: negative cytology and culture, and no clinical signs and 14 mares with endometritis: positive cytology and culture, in addition to clinical signs). Proteomic analysis was performed with a UHPLC-MS/MS system and bioinformatic analysis was carried out using Qlucore Omics Explorer. Gene Ontology enrichment and pathway analysis (PANTHER and KEGG) of the uterine proteome were performed to identify active biological pathways in enriched proteins from each group. Quantitative analysis revealed 38 proteins differentially abundant in endometritis mares when compared to healthy mares (fold changes >4.25, and q-value = 0.002). The proteins upregulated in the secretome of mares with BE were involved in biological processes related to the generation of energy and REDOX regulation and to the defense response to bacterium. A total of 24 biomarkers for BE were identified using the biomarker workbench algorithm. Some of the proteins identified were related to the innate immune system such as isoforms of histones H2A and H2B involvement in neutrophil extracellular trap (NET) formation, complement C3a, or gelsolin and profilin, two actin-binding proteins which are essential for dynamic remodeling of the actin cytoskeleton during cell migration. The other group of biomarkers were three known antimicrobial peptides (lysosome, equine cathelicidin 2 and myeloperoxidase (MPO)) and two uncharacterized proteins with a high homology with cathelicidin families. Findings in this study provide the first evidence that innate immune cells in the equine endometrium undergo reprogramming of metabolic pathways similar to the Warburg effect during activation. In addition, biomarkers of BE in uterine fluid of mares including the new proteins identified, as well as other antimicrobial peptides already known, offer future lines of research for alternative treatments to antibiotics. • Mares with bacterial endometritis exhibited a significantly higher relative abundance of endometrial proteins compared to clinically healthy mares. • The proteins upregulated in the secretome of mares with bacterial endometritis were involved in biological processes related to the generation of energy and REDOX regulation and to the defense response to bacterium (neutrophil extracellular trap (NET) formation pathway). • A total of 24 biomarkers for bacterial endometritis were identified. Some of the proteins identified were related to the innate immune system such as isoforms of histones H2A and H2B, complement C3a, or gelsolin and profilin. The other group of biomarkers were three known antimicrobial peptides (lysosome, equine cathelicidin 2 and MPO) and two uncharacterized proteins with a high homology with cathelicidin families. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are a promising alternative to antibiotics in the fight against multi-drug resistant and immune system-evading bacterial infections. Protegrins are porcine cathelicidins which have been identified in porcine leukocytes. Protegrin-1 is the best characterized family member and has broad antibacterial activity by interacting and permeabilizing bacterial membranes. Many host defense peptides (HDPs) like LL-37 or chicken cathelicidin 2 (CATH-2) have also been shown to have protective biological functions during infections. In this regard, it is interesting to study if Protegrin-1 has the immune modulating potential to suppress unnecessary immune activation by neutralizing endotoxins or by influencing the macrophage functionality in addition to its direct antimicrobial properties. This study showed that Protegrin-1 neutralized lipopolysaccharide- (LPS) and bacteria-induced activation of RAW macrophages by binding and preventing LPS from cell surface attachment. Furthermore, the peptide treatment not only inhibited bacterial phagocytosis by murine and porcine macrophages but also interfered with cell surface and intracellular bacterial survival. Lastly, Protegrin-1 pre-treatment was shown to inhibit the amastigote survival in Leishmania infected macrophages. These experiments describe an extended potential of Protegrin-1's protective role during microbial infections and add to the research towards clinical application of cationic AMPs. • Protegrin-1 neutralizes LPS- and bacteria-induced activation of RAW macrophages. • Protegrin-1 binds LPS and prevents its attachment to macrophage's cell surface. • Protegrin-1 neutralizes DPLA with more affinity than MPLA. • Bacterial phagocytosis is inhibited by Protegrin-1 treatment of murine and porcine macrophages. • Amastigote burden of Leishmania-infected RAW macrophages is lowered by Protegrin-1 treatment. [ABSTRACT FROM AUTHOR]

Cancer of the head and neck encompasses a wide range of cancers, including oral and oropharyngeal cancers. Oral cancer is often diagnosed at advanced stages and has a dismal prognosis. Piscidin-1, a marine antimicrobial peptide (AMP) containing approximately 22 amino acids, also exhibits significant anticancer properties. We investigated the possible anti-oral cancer effects of piscidin-1 and clarified the mechanisms underlying these effects. We treated the oral squamous cell carcinoma cell lines OC2 and SCC4 with piscidin-1. Cell viability and the expression of different hallmark apoptotic molecules, including reactive oxygen species (ROS), were tested using the appropriate MTT assay, flow cytometry and western blotting assays, and human umbilical vein endothelial cell (HUVEC) wound healing, migration, and tube formation (angiogenesis) assays. Piscidin-1 increases cleaved caspase 3 levels to induce apoptosis. Piscidin-1 also increases ROS levels and intensifies oxidative stress in the endoplasmic reticulum and mitochondria, causing mitochondrial dysfunction. Additionally, it decreases the oxygen consumption rates and activity of mitochondrial complexes I–V. As expected, the antioxidants MitoTEMPOL and N-acetylcysteine reduce piscidin-1–induced ROS generation and intracellular calcium accumulation. Piscidin-1 also inhibits matrix metalloproteinase (MMP)-2/-9 expression in HUVECs, affecting migration and tube formation angiogenesis. We demonstrated that piscidin-1 can promote apoptosis via both intrinsic and extrinsic apoptotic pathways and findings indicate that piscidin-1 has anti-proliferative and anti-angiogenic properties in oral cancer treatment. Our study on piscidin-1 thus provides a basis for future translational anti-oral cancer drug research and a new theoretical approach for anti-oral cancer clinical research. [Display omitted] • Piscidin-1 cleaves caspase 3 and induces apoptosis via intrinsic/extrinsic pathways. • Piscidin-1 decreased mitochondrial function, ΔΨ m , and the OXPHOS complex protein. • In OSCC cells, piscidin-1 raises intracellular Ca2+ and causes ER stress. • MitoTEMPOL and NAC mitigated the mtROS and Ca2+ caused by piscidin-1. • Piscidin-1 reduces angiogenesis in HUVECs, which is anti-tumor. [ABSTRACT FROM AUTHOR]

Owing to their limited accuracy and narrow applicability, current antimicrobial peptide (AMP) prediction models face obstacles in industrial application. To address these limitations, we developed and improved an AMP prediction model using Comparing and Optimizing Multiple DEep Learning (COMDEL) algorithms, coupled with high-throughput AMP screening method, finally reaching an accuracy of 94.8% in test and 88% in experiment verification, surpassing other state-of-the-art models. In conjunction with COMDEL, we employed the phage-assisted evolution method to screen Sortase in vivo and developed a cell-free AMP synthesis system in vitro , ultimately increasing AMPs yields to a range of 0.5–2.1 g/L within hours. Moreover, by multi-omics analysis using COMDEL, we identified Lactobacillus plantarum as the most promising candidate for AMP generation among 35 edible probiotics. Following this, we developed a microdroplet sorting approach and successfully screened three L. plantarum mutants, each showing a twofold increase in antimicrobial ability, underscoring their substantial industrial application values. This study establishes an AI-based AMPs prediction model named COMDEL for effectively mining edible AMPs and probiotics, which are subsequently synthesized and directly evolved for industrial application, respectively. [Display omitted] [ABSTRACT FROM AUTHOR]

To reduce food spoilage and deterioration caused by microbial contamination, antimicrobial peptides (AMPs) have gradually gained attention as a biological preservative. Odorranain-C1 is an α-helical cationic antimicrobial peptide extracted from the skin of frogs with broad-spectrum antimicrobial activity. In this study, we achieved the expression of Odorranain-C1 in Pichia pastoris (P. pastoris) (also known as Komagataella phaffii) by employing DNA recombination technology. The recombinant Odorranain-C1 showed broad-spectrum antibacterial activity and displayed a minimum inhibitory concentration within the range of 8–12 μg.mL-1. Meanwhile, Odorranain-C1 exhibited superior stability and lower hemolytic activity. Mechanistically, Odorranain-C1 disrupted the bacterial membrane's integrity, ultimately causing membrane rupture and subsequent cell death. In tilapia fillets preservation, Odorranain-C1 inhibited the total colony growth and pH variations, while also reducing the production of total volatile basic nitrogen (TVB-N) and thiobarbituric acid (TBA). In conclusion, these studies demonstrated the efficient recombinant expression of Odorranain-C1 in P. pastoris , highlighting its promising utilization in food preservation. • Odorranain-C1 was successfully expressed in Pichia pastoris for the first time. • Odorranain-C1 showed broad-spectrum antibacterial activity and good stability. • The mechanism of Odorranain-C1 is to destroy the integrity of bacterial membranes. • Odorranain-C1 can be used to keep tilapia fillets fresh and prolong their shelf life. [ABSTRACT FROM AUTHOR]

Treatment of subclinical mastitis (SCM) during lactation is rarely recommended due to concerns related to both antimicrobial usage and the costs associated with milk discard. Nisin is a naturally produced antimicrobial peptide with a gram-positive spectrum that, when given to dairy cows, does not require milk discard. We evaluated the economic impact of the treatment of SCM during early lactation using a nisin-based intramammary treatment under different scenarios that included various treatment costs, milk prices, and cure rates. We stochastically simulated the dynamics of SCM detected during the first week of lactation. The net economic impact was expressed in US dollars per case. The probabilities of an event and their related costs were estimated using a model that was based on pathogen-specific assumptions selected from peer-reviewed articles. Nisin cure rates were based on results of pivotal studies included in the US Food and Drug Administration (FDA) approval submission. Based on our model, the average cost of a case of intramammary infection (i.e., only true-positive cases) in early lactation was $170 (90% = $148–$187), whereas the cost of a clinical mastitis case was $521 (90% range = $435–$581). Both estimates varied with etiology, parity, and stage of lactation. When comparing the net cost of SCM cases (i.e., CMT-positive tests) detected during the first week of lactation, nisin treatment generated an average positive economic impact of $19 per CMT-positive case. The use of nisin to treat SCM was beneficial 93% of the time. Based on the sensitivity analysis, treatment would result in an economically beneficial outcome for 95% and 73% of multiparous and primiparous cows, respectively. At the herd level, use of intramammary nisin to treat SCM in cows in early lactation was economically beneficial in most tested scenarios. However, the economic impact was highly influenced by factors such as rate of bacteriological cure, cost of treatment, and parity of the affected animal. These factors should be considered when deciding to use nisin as a treatment for SCM. [ABSTRACT FROM AUTHOR]

Many factors contribute to bacterial membrane stabilization, including steric effects between lipids, membrane spontaneous curvature, and the difference in the number of neighboring molecules. This forum provides an overview of the physicochemical properties associated with membrane curvature and how this parameter can be tuned to design more effective antimicrobial peptides. [ABSTRACT FROM AUTHOR]

Ice cream manufacture commonly results in the accumulation of wasted product that contains valuable food-grade quality components, including fat, carbohydrates, and protein. Methods have been developed for recovering the fat from this waste stream, but this results in the generation of a co-product rich in fermentable carbohydrates. This study aimed to investigate the potential for using this co-product as a fermentation substrate for production of antimicrobial peptides, called bacteriocins, by dairy starter cultures. Results showed that Streptococcus thermophilus B59671 and Lactococcus lactis 11454 produced the broad-spectrum bacteriocins thermophilin 110 and nisin, respectively, when the fermentation substrate was melted ice cream, or a co-product generated by a modified butter churning technique. Bacteriocin production varied depending on the brand and variety of vanilla ice cream used in this study. When an alternate enzyme-assisted fat extraction technique was used, S. thermophilus metabolism was impaired within the resulting co-product, and thermophilin 110 production was not observed. Lactococcus lactis was still able to grow in this co-product, but antimicrobial activity was not observed. Results from this study suggest the co-product generated when using the churning technique is a better choice to use as a base medium for future studies to optimize bacteriocin production. [ABSTRACT FROM AUTHOR]

The emergence of antibiotic-resistant bacteria is a serious threat to public health. Antimicrobial peptides (AMP) are a powerful alternative to antibiotics due to their low propensity to induce bacterial resistance. However, cytotoxicity and short half-lives have limited their clinical translation. To overcome these problems, AMP conjugation has gained relevance in the biomaterials field. Nevertheless, few studies describe the influence of conjugation on enzymatic protection, mechanism of action and antimicrobial efficacy. This review addresses this gap by providing a detailed comparison between conjugated and soluble AMP. Additionally, commonly employed chemical reactions and factors to consider when promoting AMP conjugation are reviewed. The overall results suggested that AMP conjugated onto biomaterials are specifically protected from degradation by trypsin and/or pepsin. However, sometimes, their antimicrobial efficacy was reduced. Due to limited conformational freedom in conjugated AMP, compared to their soluble forms, they appear to act initially by creating small protuberances on bacterial membranes that may lead to the alteration of membrane potential and/or formation of holes, triggering cell death. Overall, AMP conjugation onto biomaterials is a promising strategy to fight infection, particularly associated to the use of medical devices. Nonetheless, some details need to be addressed before conjugated AMP reach clinical practice. Covalent conjugation of antimicrobial peptides (AMP) has been one of the most widely used strategies by bioengineers, in an attempt to not only protect AMP from proteolytic degradation, but also to prolong their residence time at the target tissue. However, an explanation for the mode of action of conjugated AMP is still lacking. This review extensively gathers works on AMP conjugation and puts forward a mechanism of action for AMP when conjugated onto biomaterials. The implications of AMP conjugation on antimicrobial activity, cytotoxicity and resistance to proteases are all discussed. A thorough review of commonly employed chemical reactions for this conjugation is also provided. Finally, details that need to be addressed for conjugated AMP to reach clinical practice are discussed. [Display omitted] [ABSTRACT FROM AUTHOR]

Animal venoms are a rich and complex source of components, including peptides (such as neurotoxins, anionic peptides and hypotensins), lipids, proteins (such as proteases, hyaluronidases and phospholipases) and inorganic compounds, which affect all biological systems of the envenoming victim. Their action may result in a wide range of clinical manifestations, including tachy/bradycardia, hyper/hypotension, disorders in blood coagulation, pain, edema, inflammation, fever, muscle paralysis, coma and even death. Scorpions are one of the most studied venomous animals in the world and interesting bioactive molecules have been isolated and identified from their venoms over the years. Tityus spp. are among the scorpions with high number of accidents reported in the Americas, especially in Brazil. Their venoms have demonstrated interesting results in the search for novel agents with antimicrobial, anti-viral, anti-parasitic, hypotensive, immunomodulation, anti-insect, antitumor and/or antinociceptive activities. Furthermore, other recent activities still under investigation include drug delivery action, design of anti-epileptic drugs, investigation of sodium channel function, treatment of erectile disfunction and priapism, improvement of scorpion antivenom and chelating molecules activity. In this scenario, this paper focuses on reviewing advances on Tityus venom components mainly through the modern omics technologies as well as addressing potential therapeutic agents from their venoms and highlighting this abundant source of pharmacologically active molecules with biotechnological application. [Display omitted] • Tityus venoms are still underexploited sources of pharmacological agents. • Only few Tityus venom sequences (of ∼3000 available at databases) have been studied in depth. • T. serrulatus venom is the most studied venom among the Tityus venoms. • Many Tityus toxins are versatile toxins presenting a broad spectrum potential. • Some Tityus venom components whose function is still unknown include the PAPE and anionic peptides. [ABSTRACT FROM AUTHOR]

Biofilm formation is usually affected by many environmental factors, including divalent cations. The purpose of the current work was to analyze how calcium (Ca2+) affects the biofilm formation of dairy Pseudomonas fluorescens isolates by investigating their growth, swarming motility, biofilm-forming capacity, extracellular polymeric substance production, and biofilm structures. Moreover, the regulation mechanism of Ca2+ involved in its biofilm formation was explored through RNA-sequencing analysis. This work revealed that supplementation of 5, 10, 15, and 20 m M Ca2+ significantly reduced the swarming motility of P. fluorescens strains (P.F 2 , P.F 4 , and P.F 17), but the biofilm-forming ability and polysaccharide production were increased after the supplementation of 5 and 10 m M Ca2+. By the supplementation of Ca2+, complex structures with more cell clusters glued together in P. fluorescens P.F 4 biofilms were confirmed by scanning electron microscopy, and increased biomass and coverage of P. fluorescens P.F 4 biofilms were observed by confocal laser scanning microscopy. In addition, RNA-sequencing results showed that P. fluorescens P.F 4 showed a transcriptional response to the supplementation of 10 m M Ca2+, and a total of 137 genes were significantly expressed. The differential genes were represented in 4 upregulated Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways (nonribosomal peptide structures, quorum sensing, biosynthesis of siderophore group nonribosomal peptides, and phenylalanine metabolism), and 4 downregulated KEGG pathways (flagellar assembly, amino sugar and nucleotide sugar metabolism, nitrotoluene degradation, and cationic antimicrobial peptide resistance). The results indicate that Ca2+ might serve as an enhancer to substantially trigger the biofilm formation of dairy P. fluorescens isolates in the dairy industry. [Display omitted] [ABSTRACT FROM AUTHOR]

Mobile colistin resistance (MCR) is the primary mechanism for colistin resistance and has been disseminated worldwide; this has gained substantial attention and led to policy changes for the use of colistin in food animals in many countries. The global spread of mcr was attributed to the high selective pressure of colistin used in livestock, and the acquisition of mcr by low-copy-number and highly conjugative plasmids. The dramatic decrease of mcr after the colistin-banning policy could be associated with the fitness cost mediated by MCR. Besides mediating resistance to both polymyxins and antimicrobial peptides, MCR is thought to enhance the virulence of bacteria and affect the host immune response. To combat colistin-resistant bacteria, MCR inhibitors have been developed based on the structure of MCR, and other strategies targeting mcr -positive plasmids/strains have also been exploited. Colistin is regarded as a last-line drug against serious infections caused by multidrug-resistant Gram-negative bacterial pathogens. Therefore, the emergence of mobile colistin resistance (mcr) genes has attracted global concern and led to policy changes for the use of colistin in food animals across many countries. Currently, the distribution, function, mechanism of action, transmission vehicles, origin of mcr , and new treatment strategies against MCR-producing pathogens have been extensively studied. Here we review the prevalence, structure and function of mcr , the fitness cost and persistence of mcr -carrying plasmids, the impact of MCR on host immune response, as well as the control strategies to combat mcr -mediated colistin resistance. Colistin is regarded as a last-line drug against serious infections caused by multidrug-resistant (MDR) Gram-negative bacterial pathogens. Therefore, the emergence of mobile colistin resistance (mcr) genes has attracted global concern and led to policy changes for the use of colistin in food animals across many countries. Currently, the distribution, function, mechanism of action, transmission vehicles, origin of mcr , and new treatment strategies against MCR-producing pathogens have been extensively studied. Here we review the prevalence, structure, and function of mcr , the fitness cost and persistence of mcr -carrying plasmids, the impact of MCR on host immune response, as well as the control strategies to combat mcr -mediated colistin resistance. [ABSTRACT FROM AUTHOR]

Bacterial biofilm-associated infection was one of the most serious threats to human health. However, effective drugs for drug-resistance bacteria or biofilms remain rarely reported. Here, we propose an innovative strategy to develop a multifunctional antimicrobial agent with broad-spectrum antibacterial activity by coupling photosensitizers (PSs) with antimicrobial peptides (AMPs). This strategy capitalizes on the ability of PSs to generate reactive oxygen species (ROS) and the membrane-targeting property of AMPs (KRWWKWIRW, a peptide screened by an artificial neural network), synergistically enhancing the antimicrobial activity. In addition, unlike conventional aggregation-caused quenching (ACQ) photosensitizers, aggregation-induced emission (AIE) PSs show stronger fluorescence emission in the aggregated state to help visualize the antibacterial mechanism. In vitro antibacterial experiments demonstrated the excellent killing effects of the developed agent against both Gram-positive (G+) and Gram-negative (G–) bacteria. The bacterial-aggregations induced ability enhanced the photoactivatable antibacterial activity against G– bacteria. Notably, it exhibited a significant effect on destroying MRSA biofilms. Moreover, it also showed remarkable efficacy in treating wound infections in mice in vivo. This multifunctional antimicrobial agent holds significant potential in addressing the challenges posed by bacterial biofilm-associated infections and drug-resistant bacteria. Schematic illustration of TPI-CysHHC10 comprising AMPs and AIE photosensitizer as a photoactivatable, broad-spectrum, antibacterial agent for combating biofilms. [Display omitted] [ABSTRACT FROM AUTHOR]

Due to the emergence of drug resistance of pathogens, it seem various strategies to be required to discover and develop new antimicrobial agents. Plant antimicrobial peptides (AMPs) can be promising alternative candidates against a variety of pathogens. Recently, various plant peptides have been identified and classified. Plant defensins are a class of cysteine-rich peptides with broad antimicrobial spectrum effects. In this study, an antimicrobial plant defensin peptide was identified successfully from EST library of garlic (Allium sativum L.) as well as expressed in E. coli expression system. The coding sequence of garlic defensin consist of an open reading frame (ORF) containing 216 base pairs that encodes a peptide of 71 amino acids with a putative extracellular signal peptide (24 amino acids) at the N- terminus. Analyze experimental data obtained from antimicrobial susceptibility assays shows that the garlic defensin has antibacterial effect against Escherichia coli and Staphylococcus aureus as models for Gram-negative and Gram positive bacteria, respectively. On the other hand, it was found that this defensin also possessed temperature and pH stability and its activity remained constant in different temperature and pH conditions. In summary, our findings provides important insights for producing and developing new potential antimicrobial compounds by using chemical synthesis and/or recombinant techniques in the future. [Display omitted] • Antimicrobial peptides represent a large family of small cysteine-rich cationic peptides. • Plant defensins play important roles in plant immune responses against pathogenic bacteria. • A plant defensin from garlic (Allium sativum L.) were identified using in silico and in vitro methods. • Some structural, physicochemical, and biological features of identified defensin were investigated and characterized. • This defensin appears can be considered as a potential peptide antibiotic drug for development and further assessment in the future. [ABSTRACT FROM AUTHOR]

Fungal keratitis is the foremost cause of corneal infections worldwide, of which Fusarium spp. is the common etiological agent that causes loss of vision and warrants surgical intervention. An increase in resistance to the available drugs along with severe side effects of the existing antifungals demands for new effective antimycotics. Here, we demonstrate that antimicrobial peptide S100A12 directly binds to the phospholipids of the fungal membrane, disrupts the structural integrity, and induces generation of reactive oxygen species in fungus. In addition, it inhibits biofilm formation by Fusarium spp. and exhibits antifungal property against Fusarium spp. both in vitro and in vivo. Taken together, our results delve into specific effect of S100A12 against Fusarium spp. with an aim to investigate new antifungal compounds to combat fungal keratitis. [ABSTRACT FROM AUTHOR]

Plant-derived peptides represent a promising group of natural compounds with broad industrial and pharmaceutical applications. Low-efficiency production level is the major obstacle to the commercial production of such bioactive peptides. Today, recombinant techniques have been developed for fast and cost-effective production of high-quality peptides for various applications in the chemical and food industries. The roseltide rT1 is a plant peptide with different antimicrobial properties and therapeutic applications in the prevention and treatment of inflammatory lung diseases by inhibiting human neutrophil elastases. Here, we report the expression of functional recombinant roseltide rT1 peptide in tobacco plants. Transgenic plants were generated by the Agrobacterium-mediated transformation method followed by molecular analysis of transgenic plants to demonstrate successful integration and expression of recombinant rT1 peptide. Protein extracts of transgenic plants expressing a single-copy rT1 gene showed efficient antimicrobial properties as verified by growth inhibition of different bacterial strains. Our results illustrate that plant-derived recombinant rT1 peptide is a promising alternative for rapid and cost-effective production of this important antimicrobial peptide for application in therapeutic and food industries. • Transgenic tobacco plants expressing recombinant roseltide rT1 were generated by Agrobacterium-mediated transformation. • Single-copy transgenic lines were selected and verified by different molecular methods. • The expressed recombinant peptide performed efficient antibacterial properties. • Plant system is presented as a promising alternative for rapid and cost-effective production of recombinant rT1 peptide. [ABSTRACT FROM AUTHOR]

The antimicrobial peptide (AMP) LI is a fusion product of antimicrobial peptide LL37 produced by human neutrophils and Indolicidin secreted by bovine neutrophils. LI retained the antimicrobial activity of the parental peptides and showed high cell selectivity. In this study, the flexible linker Gly-Ser-Gly (G-S-G) was used to ligate LI into dimeric LIG, and constructed the Pichia pastoris (P. pastoris) expression vector pPIC9K-6×His-3×FLAG-LIG. The total protein expression of P. pastoris GS115 reached the highest level (189.6 mg/L) after 96 h induction with 3 % methanol at the initial pH value of 7.0. Finally, 5.9 mg/L of recombinant LIG (rLIG) was obtained after enterokinase digestion and purification. The rLIG had high antimicrobial activity and low hemolytic activity. Compared with monomer LI, GSG linked dimeric LIG, which had no significant change in antimicrobial activity and had good salt ions stability. In this study, the dimeric antimicrobial peptide LIG was successfully expressed, which provided a new idea for the expression of AMPs in the P. pastoris expression system, and had important significance for the application of AMPs. • The flexible linker GSG was used to construct the dimeric antimicrobial peptide LIG. • The antimicrobial peptide LIG was successfully expressed by Pichia pastoris. • The recombinant antimicrobial peptide LIG has high biological activity. [ABSTRACT FROM AUTHOR]

Antibiotic abuses cause pathogen resistance to conventional antibiotics, threating public health globally. Antimicrobial peptides were considered as substitutes for antibiotic because of their broaden and rapid antibacterial activity. BmKbpp extracted from the Chinese scorpion Mesobuthus martensii Karsch inhibits the growth of gram-negative bacteria and fungi efficiently, while its efficacy for inhibiting the growth of the gram-positive Staphylococcus aureus was limited. Besides, how BmKbpp inhibits bacterial growth remains elusive thus far. In this study, a C-terminal hemagglutinin (HA) and 6×His (HA-6×His) tagged BmKbpp of three repeats (3×BmKbpp) was expressed in Chlamydomonas reinhardtii. The recombinant 3×BmKbpp-HA-6×His exhibited antibacterial activity against both gram-positive and gram-negative bacteria with a minimum inhibitory concentration between 20⁓200 µg/ml. The minimum bactericidal concentration ranged from 1 × MIC to 3 × MIC for 3×BmKbpp-HA-6×His. 3×BmKbpp-HA-6×His exhibited a good thermostability and resisted on pH shock and digestion of certain proteinases. It also displayed low cytotoxicity and hemolysis as tested on cell lines including HEK293, Vero, MDCK, and BHK-21. In addition, 3×BmKbpp-HA-6×His inhibits bacterial growth by effectively distorting bacterial cell membrane. Thus, C. reinhardtii -derived 3×BmKbpp-HA-6×His effectively inhibits bacterial growth, making it a promising antimicrobial peptide. [Display omitted] • A triple BmKbpp (3×BmKbpp) was stably expressed in Chlamydomonas reinhardtii. • 3×BmKbpp inhibits the growth of a wide spectrum of bacteria. • 3×BmKbpp was thermostable and tolerant of pH shock and certain proteinases. • 3×BmKbpp showed a low toxicity to mammalian and human cells. • 3×BmKbpp disrupted the bacterial cell membrane integrity. [ABSTRACT FROM AUTHOR]

The antimicrobial peptides SAAP-148 and Ab-Cath were encapsulated in oleyl-modified hyaluronic acid (OL-HA) nanogels to improve these peptides' selectivity index. The physicochemical properties, antimicrobial activity, and cytotoxicity of the AMP-loaded OL-HA nanogels were studied in vitro. Results showed good physicochemical properties of the nanogels; polydisperse nanogels (PDI=0.2) ranging in size from 181 to 206 nm, with high negative surface charge (−47 to −48 mV) and moderate encapsulation efficiency (EE=53–63%). Encapsulation of SAAP-148 or Ab-Cath in nanogels maintained antimicrobial activity against antimicrobial resistant Staphylococcus aureus and Acinetobacter baumannii , while reducing the peptides' hemolytic activity against human erythrocytes and cytotoxicity against human primary skin fibroblasts. Thus, encapsulation in OL-HA nanogels improved the overall selectivity index of SAAP-148 and Ab-Cath and their potential as therapeutics to combat AMR bacterial infections. [Display omitted] Antimicrobial peptides (AMPs) are promising alternatives to antibiotics for treatment of antimicrobial resistant (AMR) bacterial infections. However, their narrow therapeutic window due to in vivo toxicity and limited stability hampers their clinical use. Here, we evaluated encapsulation of two amphiphilic AMPs, SAAP-148 and snake cathelicidin Ab-Cath, into oleyl-modified hyaluronic acid (OL-HA) nanogels to improve their selectivity index. The AMP-loaded OL-HA nanogels ranged 181–206 nm in size with a PDI of 0.2, highly negative surface charge (−47 to −48 mV) and moderate encapsulation efficiency (53–63%). The AMP-loaded OL-HA nanogels displayed similar activity in vitro as AMP solutions against AMR Staphylococcus aureus and Acinetobacter baumannii , with a dose-dependent effect over time. Importantly, the AMP-loaded OL-HA nanogels showed decreased cytotoxicity towards human erythrocytes and primary skin fibroblast, thereby improving the selectivity index of SAAP-148 and Ab-Cath by 2- and 16.8-fold, respectively. Particularly, the selectivity of Ab-Cath-loaded OL-HA nanogels has great clinical potential, with an index that reached ≥ 300 for S. aureus and ≥ 3000 for A. baumannii. These findings indicate that OL-HA nanogels are a promising drug delivery system to reduce the cytotoxicity of AMPs without substantially affecting their antimicrobial activity, thereby increasing their selectivity index and potential as therapeutics to combat AMR bacterial infections. [ABSTRACT FROM AUTHOR]

Hepcidin has been identified as an important antimicrobial peptide exerting important innate immunomodulatory activities in many fish species. In the present study, reverse transcription PCR coupled with the rapid amplification of cDNA ends was used to obtain the full-length cDNA of the crescent sweetlips hepcidin gene, which is 829 bp in length and includes an 273 bp ORF encoding a peptide with 90 amino acid residues. Sequence alignment showed a typical RXKR motif and eight conserved cysteine residues in the deduced amino acid sequences. Four disulfide bonds were predicted to form between these eight cysteines, which may stabilize the hairpin structure in hepcidin molecule. Furthermore, phylogenetic analysis showed that the deduced amino acid sequences of crescent sweetlips hepcidin had high sequence homology to hepcidins from fish species of Eupercaria. In addition, the crescent sweetlips hepcidin peptide demonstrated a strong antimicrobial activity in vitro against several types of pathogenic bacteria in fish. In conclusion, the obtained results suggested that crescent sweetlips hepcidin possessed the typical structure similar to other fish hepcidins and had strong antibacterial activity, which showed great potential in the prevention of fish diseases in aquaculture. • Hepcidin full-length cDNA sequence was cloned from a marine fish crescent sweetlips. • Crescent sweetlips hepcidin peptide exhibited desirable antibacterial activity. • Disulfide bonds between eight cysteines were crucial for the antibacterial activity. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are essential for defence against pathogens in all living organisms and possessed activities against bacteria, fungi, viruses, parasites and even cancer cells. AMPs are short peptides containing 12–100 amino acids conferring a net positive charge and an amphiphilic property in most cases. Although, anionic AMPs also exist. AMPs can be classified based on the types of secondary structures, charge, hydrophobicity, amino acid composition, length, etc. Their mechanism of action usually includes a membrane disruption process through pore formation (three different models have been described, barrel-stave, toroidal or carpet model) but AMPs can also penetrate and impair intracellular functions. Besides their activity against pathogens, they have also shown immunomodulatory properties in complex scenarios through many different interactions. The aim of this review to summarize knowledge about AMP's and discuss the potential application of AMPs as therapeutics, the challenges due to their limitations, including their susceptibility to degradation, the potential generation of AMP resistance, cost, etc. We also discuss the current FDA-approved drugs based on AMPs and strategies to circumvent natural AMPs' limitations. • Antimicrobial peptides (AMPs) are essential for defence against pathogens. • AMPs possessed activities against bacteria, fungi, viruses, parasites and even cancer cells. • AMPs are short peptides containing 12-100 amino acids conferring a net positive charge and an amphiphilic property. • AMPs have shown immunomodulatory properties in complex scenarios through many different interactions. [ABSTRACT FROM AUTHOR]

The power generation and pollutants removal efficiencies of mixed electroactive microbial community rely on the effective electron transfer between outer membrane proteins of electroactive electricigens. However, the poor conductivity of protein and various non-electrogenic bacteria greatly hinder the electron transfer and effective contact between electricigens. Herein, we report a novel "bi-directional" microbial community regulation method that can selectively inhibit Gram-positive non-electrogenic bacteria and enhances the conductivity and electron transfer between electrogenic bacteria and the electrode in MFC by taking the advantages of the excellent conductive and bacteriostatic properties of Au/Nisin nanocomposite. The Au/Nisin stimulation endows microbial fuel cell (MFC) a high-efficiency functional community, which increases the power density and decolorization rate by 103 % and 33 %, respectively. The high conductivity, and selectivity and antimicrobial activity of the nanocomposite improve the electrochemical activity of the bioanode, hereby promoting the microbial extracellular electron transfer and increasing the proportion of electrogenic bacteria such as Proteobacteria , Bacteroidetes and Desulfovibrio in the biofilm. Our work successfully demonstrates that the construction of multifunctional metallic peptide nanocomposite can effectively regulate microbial community to improve the electrochemical performance of MFC. [Display omitted] • Antibacterial and conductive Au/Nisin was prepared as microbial community regulator. • Au/Nisin significantly improves the electricity generation and decolorization of MFC. • A novel strategy was provided to realize bidirectional microbial community regulation. [ABSTRACT FROM AUTHOR]

Human scalp hair follicles (HFs) engage in olfactory receptor (OR)-dependent chemosensation. Activation of olfactory receptor family 2 subfamily AT member 4 (OR2AT4) by the synthetic, sandalwood-like odorant Sandalore® up-regulated HF antimicrobial peptide expression of dermcidin (DCD), which had previously been thought to be produced exclusively by sweat and sebaceous glands. To understand if intrafollicular DCD production can be stimulated by a commonly used cosmetic odorant, thus altering human HF microbiome composition in a clinically beneficial manner. DCD expression was compared between fresh-frozen scalp biopsies and microdissected, full-length scalp HFs, organ-cultured in the presence/absence of the OR2AT4 agonist, Sandalore® and/or antibiotics and/or the competitive OR2AT4 antagonist, Phenirat®. Amplicon-based sequencing and microbial growth assays were performed to assess how this treatment affected the HF microbiome. Synthetic odorant treatment upregulated epithelial DCD expression and exerted antimicrobial activity in human HFs ex vivo. Combined antibiotic and odorant treatment, during an ex vivo dysbiosis event, prevented HF tissue damage and favoured a more physiological microbiome composition. Sandalore®-conditioned medium, containing higher DCD content, favoured Staphylococcus epidermidis and Malassezia restricta over S. aureus and M. globosa, while exhibiting antimicrobial activity against Cutibacterium acnes. These effects were reversed by co-administration of Phenirat®. We provide the first proof-of-principle that a cosmetic odorant impacts the human HF microbiome by up-regulating antimicrobial peptide production in an olfactory receptor-dependent manner. Specifically, a synthetic sandalwood-like odorant stimulates intrafollicular DCD production, likely via OR2AT4, and thereby controls microbial overgrowth. Thus, deserving further exploration as an adjuvant therapeutic principle in the management of folliculitis and dysbiosis-associated hair diseases. • A synthetic odorant upregulates intrafollicular antimicrobial peptide expression. • Under dysbiotic conditions, odorant/antibiotic co-treatment prevents hair follicle damage. • Antibiotic and synthetic odorant treatment favours a more physiological microbiome. • Odorant-induced dermcidin production regulates Staphylococcus and Malassezia. • Odorant effects on follicular microbiome are olfactory receptor 2AT4-mediated. [ABSTRACT FROM AUTHOR]

The production of antimicrobial peptides/proteins (AMPs) in sufficient quantities for clinical evaluation is challenging because complex peptides are unsuitable for chemical synthesis, natural sources have low yields, and heterologous systems often have low expression levels or require product-specific process adaptations. Here we describe the production of a complex AMP, the insect metalloproteinase inhibitor (IMPI), by adding a C-terminal C-tag to increase the yield compared to the unmodified peptide. We used a design of experiments approach for process intensification in Escherichia coli Rosetta-gami 2(DE3)pLysS cells and achieved a yield of 260 mg L−1, which is up to 30-fold higher than previously reported. The C-tag also enhanced product purity but had no effect on IMPI activity, making tag removal unnecessary and therefore simplifying process analytics and downstream processing. We have confirmed that the C-tag is compatible with the peptide and could form the basis of a platform technology for the expression, purification and detection of diverse AMPs produced in E. coli. [ABSTRACT FROM AUTHOR]

Bacterial infections and excessive inflammation can impede the healing of wounds. Hydrogels have emerged as a promising approach for dressing bacterial-infected injuries. However, some antibacterial hydrogels are complex, costly, and even require assistance with other instruments such as light, making them unsuitable for routine outdoor injuries. Here, we developed an in-situ generating hydrogel via hybridizing oxidized β -D-glucan with antimicrobial peptide C 8 G 2 through the Schiff base reaction. This hydrogel is easily accessible and actively contributes to the whole healing process of bacterial-infected wounds, demonstrating remarkable antibacterial activity and biological compatibility. The pH-sensitive reversible imine bond enables the hydrogel to self-heal and sustainably release the antibacterial peptide, thereby improving its bioavailability and reducing toxicity. Meanwhile, the immunoregulating β -D-glucan inhibits the release of inflammatory factors while promoting the release of anti-inflammatory factors. In methicillin-resistant Staphylococcus aureus (MRSA)-infected full-thickness skin wound models, the hybrid hydrogel showed superior antibacterial and anti-inflammatory activity, enhanced the M2 macrophage polarization, expedited wound closure, and regenerated epidermis tissue. These features make this hydrogel an appealing wound dressing for treating multi-drug-resistant bacteria-infected wounds. [Display omitted] • The BGA/C 8 G 2 injectable hydrogel can form rapidly in situ. • The hydrogel releases the antibacterial peptide in acidic bacterial infections. • The hydrogel can participate in the entire wound healing process. • The hydrogel accelerates MRSA-infected wound healing without additions. [ABSTRACT FROM AUTHOR]

[Display omitted] • Pretreatment with AKK or pAKK modulates S. Typhimurium infection. • AKK enhances the gut barrier and increases RegIII lectins secretion after infection. • pAKK stimulates antimicrobial activity, ROS production, and inflammasome activation in macrophages and mice. • AKK-altered gut microbiota plays a partial role in attenuating infection. The gut microbiome is vital for providing resistance against colonized pathogenic bacteria. Recently, specific commensal species have become recognized as important mediators of host defense against microbial infection by a variety of mechanisms. To examine the contribution of live and pasteurized A. muciniphila to defend against the intestinal pathogen Salmonella Typhimurium in a streptomycin-treated mouse model of infection. C57B6J mice were pretreated with phosphate-buffered saline (PBS), live Akkermansia muciniphila (AKK), and pasteurized A. muciniphila (pAKK) for two weeks, then mice were infected by S. Typhimurium SL 1344. 16S rRNA-based gut microbiota analysis was performed before and after infection. Bacterial counts in feces and tissues, histopathological analysis, gut barrier-related gene expression, and antimicrobial peptides were examined. Co-housing was performed to examine the role of microbiota in the change of susceptibility of mice to infection. AKK and pAKK markedly decrease d Salmonella fecal and systemic burdens and reduced inflammation during infection. Notably, further characterization of AKK and pAKK protective mechanisms revealed different candidate protective pathways. AKK promoted gut barrier gene expression and the secretion of antimicrobial peptides, and co-housing studies suggested that AKK-associated microbial community played a role in attenuating infection. Moreover, pAKK had a positive effect on NLRP3 in infected mice. We verified that pretreatment of pAKK could promote the expression of NLRP3, and enhance the antimicrobial activity of macrophage, likely through increasing the production of reactive oxygen (ROS), nitric oxide (NO), and inflammatory cytokines. Our study demonstrates that live or pasteurized A. muciniphila can be effective preventive measures for alleviating S. Typhimurium-induced disease, highlighting the potential of developing Akkermansia-based probiotics or postbiotics for the prevention of Salmonellosis. [ABSTRACT FROM AUTHOR]

Urinary tract infections (UTIs) are among the most common infections in humans. Current challenges include increasing antimicrobial resistance, frequent recurrence, and lack of vaccines or adjunctive immunotherapies. Recent work, mostly in preclinical models, has unveiled new paradigms in urinary tract immune defenses. These include advances in understanding how myeloid cells modulate UTI susceptibility and outcomes, and a new phagocytic role for collecting duct intercalated cells. Advances in our mechanistic understanding of the host–pathogen conversation can help shape future immunomodulatory therapies for UTIs. Epithelial remodeling within the bladder after uropathogenic Escherichia coli (UPEC) infection underlies increased susceptibility to recurrent UTI. During cystitis, specific populations of bladder tissue-resident macrophages and memory T cells mediate complex responses to acute and recurrent UTI. A sex-dependent defect in neutrophil maturation within the kidney contributes to the increased severity of pyelonephritis seen in androgen-exposed hosts. Complement factor C5a, acting through its receptors C5aR1 and C5aR2, enhances renal proinflammatory responses and promotes UPEC colonization of the renal tubular space. Intercalated cells within the renal collecting duct represent nonprofessional phagocytes, taking up uropathogenic bacteria and secreting antimicrobial peptides. Recent advances in preclinical modeling of urinary tract infections (UTIs) have enabled the identification of key facets of the host response that influence pathogen clearance and tissue damage. Here, we review new insights into the functions of neutrophils, macrophages, and antimicrobial peptides in innate control of uropathogens and in mammalian infection-related tissue injury and repair. We also discuss novel functions for renal epithelial cells in innate antimicrobial defense. In addition, epigenetic modifications during bacterial cystitis have been implicated in bladder remodeling, conveying susceptibility to recurrent UTI. In total, contemporary work in this arena has better defined host processes that shape UTI susceptibility and severity and might inform the development of novel preventive and therapeutic approaches for acute and recurrent UTI. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are promising anti-infective drugs, but their use is restricted by their short-term retention at the infection site, non-targeted uptake, and adverse effects on normal tissues. Since infection often follows an injury (e.g. , in a wound bed), directly immobilizing AMPs to the damaged collagenous matrix of the injured tissues may help overcome these limitations by transforming the extracellular matrix microenvironment of the infection site into a natural reservoir of AMPs for sustained in situ release. Here, we developed and demonstrated an AMP-delivery strategy by conjugating a dimeric construct of AMP Feleucin-K3 (Flc) and a collagen hybridizing peptide (CHP), which enabled selective and prolonged anchoring of the Flc-CHP conjugate to the damaged and denatured collagen in the infected wounds in vitro and in vivo. We found that the dimeric Flc and CHP conjugate design preserved the potent and broad-spectrum antimicrobial activities of Flc while significantly enhancing and extending its antimicrobial efficacy in vivo and facilitating tissue repair in a rat wound healing model. Because collagen damage is ubiquitous in almost all injuries and infections, our strategy of targeting collagen damage may open up new avenues for antimicrobial treatments in a range of infected tissues. The concept of in situ protection against bacterial infection enabled by immobilizing an antimicrobial peptide to the injured tissue through conjugation with a collagen hybridizing peptide. [Display omitted] [ABSTRACT FROM AUTHOR]

Salmonoid cathelicidin (termed CATH_BRALE) is a fish-derived antimicrobial peptide (AMP) isolated from Brachymystax lenok. In this study, CATH_BRALE was expressed in Chlamydomonas reinhardtii as a format of three repeats (3×CATH_BRALE) attached with hemagglutinin (HA) and 6×His at its C-terminus, producing a recombinant peptide with a molecular weight of ∼18 kDa. The recombinant 3×CATH_BRALE-HA-6×His remains to be expressed stably in Chlamydomonas cells even after passaging continuously for five months and yields up to 0.21% of the total Chlamydomonas soluble proteins. 3×CATH_BRALE-HA-6×His showed broad-spectrum antibacterial activity against bacteria, with MIC values ranging from 40 to 50 µg/ml for Gram-negative bacteria and 30–40 µg/ml for Gram-positive bacteria. This recombinant peptide had strong thermostability and pH stability, and its antibacterial activity was rarely altered when temperature and pH are changed. It resisted to the digestion of several tested proteases to certain extents. Besides, 3×CATH_BRALE-HA-6×His is biologically safe as it did not hemolyze rat erythrocytes nor caused cytotoxicity on Vero, BHK21, HEK293, and MDBK cells. Its antibacterial action was achieved by penetrating the cell membrane to disrupt the membrane of the target bacterial cell. In sum, our data showed that C. reinhardtii can be used as a heterologous expression host to produce biologically active CATH_BRALE. [Display omitted] • A multimer termed 3 ×CATH_BRALE was stably expressed in Chlamydomonas reinhardtii. • C. reinhardtii-derived 3 ×CATH_BRALE showed a wide spectrum of antibacterial activity. • C. reinhardtii-derived 3 ×CATH_BRALE was tolerant of temperature, pH, and proteinases. • C. reinhardtii-derived 3 ×CATH_BRALE was not toxic to mammalian and human cells. • C. reinhardtii-derived 3 ×CATH_BRALE permeabilizes the bacterial cell membrane. [ABSTRACT FROM AUTHOR]

The antimicrobial peptides Mytichitin-CB and Hispidalin were isolated from Mytilus coruscus and Benincasa hispida , respectively. Our previous studies showed that Mytichitin-CB, when expressed in Chlamydomonas reinhardtii , inhibits the bacterial growth with moderate efficiency. Naturally isolated or Pichia pastoris -expressed Hispidalin also exerts moderate efficiency for inhibiting the growth of bacteria. Here, we expressed a chimeric peptide composed of Mytichitin-CB and Hispidalin that we named as MCH in C. reinhardtii. MCH was stably expressed and its yield accounted for 0.18% of total soluble proteins of the host cells. C. reinhardtii -derived MCH inhibited the growth of both Gram-positive and Gram-negative bacteria by disrupting their cell membrane integrity. Its minimal inhibitory concentration (MIC) were ranged between 20 and 40 μg/mL, demonstrating that MCH is more potent for inhibiting the bacterial growth than individual microalgae-expressed Mytichitin-CB and yeast-expressed Hispidalin. Of more importance, MCH was temperature- and pH-tolerant, protease-resistant and exhibited low hemolytic activity against rat erythrocytes and low cytotoxicity to human embryonic kidney 293 (HEK 293) and Madin-Darby bovine kidney (MDBK) cells. Therefore, we conclude that C. reinhardtii -derived chimeric peptide MCH has a great potential for being developed to replace traditional antibiotics in the future. [Display omitted] • Chimeric MCH peptide was stably expressed in Chlamydomonas reinhardtii. • C. reinhardtii -derived MCH displayed enhanced antibacterial activity. • C. reinhardtii -derived MCH was tolerant of temperature, pH, and proteinases. • C. reinhardtii -derived MCH was not toxic to mammalian and human cells. • C. reinhardtii -derived MCH disrupted membrane integrity of bacteria. [ABSTRACT FROM AUTHOR]

The increase in antibiotic resistance calls for accelerated molecular engineering strategies to diversify natural products for drug discovery. The incorporation of non-canonical amino acids (ncAAs) is an elegant strategy for this purpose, offering a diverse pool of building blocks to introduce desired properties into antimicrobial lanthipeptides. We here report an expression system using Lactococcus lactis as a host for non-canonical amino acid incorporation with high efficiency and yield. We show that incorporating the more hydrophobic analog ethionine (instead of methionine) into nisin improves its bioactivity against several Gram-positive strains we tested. New-to-nature variants were further created by click chemistry. By azidohomoalanine (Aha) incorporation and subsequent click chemistry, we obtained lipidated variants at different positions in nisin or in truncated nisin variants. Some of them show improved bioactivity and specificity against several pathogenic bacterial strains. These results highlight the ability of this methodology for lanthipeptide multi-site lipidation, to create new-to-nature antimicrobial products with diverse features, and extend the toolbox for (lanthi)peptide drug improvement and discovery. [ABSTRACT FROM AUTHOR]

Recently, much emphasis has been placed on solving the intrinsic defects of antimicrobial peptides (AMPs), especially their susceptibility to protease digestion for the systemic application of antibacterial biomaterials. Although many strategies have increased the protease stability of AMPs, antimicrobial activity was severely compromised, thereby substantially weakening their therapeutic effect. To address this issue, we introduced hydrophobic group modifications at the N-terminus of proteolysis-resistant AMPs D1 (AArIIlrWrFR) through end-tagging with stretches of natural amino acids (W and I), unnatural amino acid (Nal) and fatty acids. Of these peptides, N1 tagged with a Nal at N-terminus showed the highest selectivity index (GM SI =19.59), with a 6.73-fold improvement over D1. In addition to potent broad-spectrum antimicrobial activity, N1 also exhibited high antimicrobial stability toward salts, serum and proteases in vitro and ideal biocompatibility and therapeutic efficacy in vivo. Furthermore, N1 killed bacteria through multiple mechanisms, involving disruption of bacterial membranes and inhibition of bacterial energy metabolism. Indeed, appropriate terminal hydrophobicity modification opens up new avenues for developing and applying high-stability peptide-based antibacterial biomaterials. To improve the potency and stability of proteolysis-resistant antimicrobial peptides (AMPs) without increasing toxicity, we constructed a convenient and tunable platform based on different compositions and lengths of hydrophobic end modifications. By tagging an Nal at the N-terminal, the obtained target compound N1 exhibited strong antimicrobial activity and desirable stability under multifarious environments in vitro (proteases, salts and serum), and also showed favorable biocompatibility and therapeutic efficacy in viv o. Notably, N1 exerted its bactericidal effect by damaging bacterial cell membranes and inhibiting bacterial energy metabolism in a dual mode. The findings provide a potential method for designing or optimizing proteolysis-resistant AMPs thus promoting the development and application of peptide-based antibacterial biomaterial. [Display omitted] [ABSTRACT FROM AUTHOR]

Recently, liver-expressed antimicrobial peptide 2 (LEAP2) was identified as an endogenous antagonist and an inverse agonist of the ghrelin receptor GHSR. However, its functions in lower vertebrates are not well understood. Our recent study demonstrated that both LEAP2 and ghrelin are functional towards a fish GHSR from Latimeria chalumnae , an extant coelacanth believed to be one of the closest ancestors of tetrapods. However, amino acid sequence alignment identified that the 6.58 position (Ballesteros-Weinstein numbering system) of most fish GHSRs are not occupied by an aromatic Phe residue, which is absolutely conserved in all known GHSRs from amphibians to mammals, and is responsible for human GHSR binding to its agonist, ghrelin. To test whether these unusual fish receptors are functional, we studied the ligand binding properties of three representative fish GHSRs, two from Danio rerio (zebrafish) and one from Larimichthys crocea (large yellow croaker). After overexpression in human embryonic kidney 293T cells, the three fish GHSRs retained normal binding to all tested LEAP2s, except for a second LEAP2 from L. crocea. However, they displayed almost no binding to all chemically synthesized n -octanoylated ghrelins, despite these ghrelins all retaining normal function towards human and coelacanth GHSRs. Thus, it seems that LEAP2 is a more conserved ligand than ghrelin towards fish GHSRs. Our results not only provided new insights into the interaction mechanism of GHSRs with LEAP2s and ghrelins, but also shed new light on the functions of LEAP2 and ghrelin in different fish species. [Display omitted] • Ghrelin, LEAP2, and their receptor GHSR are highly conserved in all vertebrates. • Most fish GHSRs lack the highly conserved Phe6.58, raising doubts for their function. • Fish GHSRs from D. rerio and L. crocea almost lost binding to all tested ghrelins. • However, these unusual fish GHSRs bound to all LEAP2s, except LEAP2B from L. crocea. • LEAP2 is a more conserved ligand than ghrelin towards fish GHSRs. [ABSTRACT FROM AUTHOR]

Systematic structural studies of gap junctions, host defense peptides, importers, and exporters have provided insights into channel stoichiometries and topologies, pore diameters, and functional states that mediate second messenger 2′3′-cGAMP and cyclic dinucleotide (CDN) transport. Comprehensive structural investigation of a range of 2′3′-cGAMP transporters has outlined the diversity of intracellular, membrane, and extracellular components that constitute channel architecture, thereby defining channels with distinct functional capabilities. The structures of CDNs including 2′3′-cGAMP bound to the SLC19A1 channel have highlighted the intertwined compact dimeric second messenger fold and its positioning in an inward-open channel, thereby opening opportunities for investigation of the corresponding outward-open and occluded states towards a molecular understanding of the transport cycle. The positioning of monomeric folates and antifolates in the interior of the SLC19A1 channel has provided a foundation for future design and optimization of agonists and antagonists impacting on 2′3′-cGAMP transport, with therapeutic implications for inflammatory and immune-associated pathologies. Recent studies have provided insights into the regulated channel-mediated transport of 2′3′-cGAMP associated with the cGAS–STING signaling pathway in inflammation from a structural and functional perspective. This has defined an underlying mechanism for 2′3′-cGAMP as an 'immunotransmitter' that mediates intercellular communication. Structures are available for apo-, 2′3′-cGAMP-, folate-, and antifolate-bound states of the SLC19A1 channel, and these provide insights into both the transport cycle and approaches for generating and testing small-molecule agonists and antagonists that impact on second messenger transport. Upon activation by double-stranded DNA (dsDNA), the cytosolic dsDNA sensor cyclic GMP-AMP synthase (cGAS) synthesizes the diffusible cyclic dinucleotide 2′3′-cGAMP (cyclic GMP-AMP), which subsequently binds to the adaptor STING, triggering a cascade of events leading to an inflammatory response. Recent studies have highlighted the role of 2′3′-cGAMP as an 'immunotransmitter' between cells, a process facilitated by gap junctions as well as by specialized membrane-spanning importer and exporter channels. This review highlights recent advances from a structural perspective of intercellular trafficking of 2′3′-cGAMP, with particular emphasis on the binding of importer SLC19A1 to 2′3′-cGAMP, as well as the significance of associated folate nutrients and antifolate therapeutics. This provides a path forward for structure-guided understanding of the transport cycle in immunology, as well as for candidate targeting approaches towards therapeutic intervention in inflammation. [ABSTRACT FROM AUTHOR]

[Display omitted] • M. anisopliae phenylethyl alcohol exerts an avoidance effect to locusts. • M. anisopliae infection elevates the expression of LmOBP11 in locusts. • Treatment with phenylethyl alcohol upregulates LmOBP11 in locusts. • Locust LmOBP11 is involved in the detection of fungal phenylethyl alcohol. • LmOBP11 negatively regulates the locust immune system to benefit fungal infection. Odorant-binding proteins (OBPs) are a class of small molecular weight soluble proteins that exist as expanded gene families in all insects, acting as ligand carriers mediating olfaction and other physiological processes. During fungal infection, a subset of insect OBPs were shown to be differentially expressed. We tested whether the altered expression of insect OBP s during pathogenic infection plays a role in behavioral or immune interactions between insect hosts and their pathogens. A wide range of techniques including RNAi-directed knockdown, heterologous protein expression, electrophysiological/behavioral analyses, transcriptomics, gut microbiome analyses, coupled with tandem mass spectrometry ion monitoring, were used to characterize the function of a locust OBP in host behavioral and immune responses. The entomopathogenic fungus Metarhizium anisopliae produces the volatile compound phenylethyl alcohol (PEA) that causes behavioral avoidance in locusts. This is mediated by the locust odorant binding protein 11 (LmOBP11). Expression of LmOBP11 is induced by M. anisopliae infection and PEA treatment. LmOBP11 participates in insect detection of the fungal-produced PEA and avoidance of PEA-contaminated food, but the upregulation of LmOBP11 upon M. anisopliae infection negatively affects the insect immune responses to ultimately benefit successful mycosis by the pathogen. RNAi knockdown of LmOBP11 increases the production of antimicrobial peptides and enhances locust resistance to M. anisopliae infection, while reducing host antennal electrophysiological responses to PEA and locust avoidance of PEA treated food. Also, transcriptomic and gut microbiome analyses reveal microbiome dysbiosis and changes in host genes involved in behavior and immunity. These results are consistent with the elevated expression of LmOBP11 leading to enhanced volatile detection and suppression of immune responses. These findings suggest a crosstalk between olfaction and immunity, indicating manipulation of host OBPs as a novel target exploited by fungal pathogens to alter immune activation and thus promote the successful infection of the host. [ABSTRACT FROM AUTHOR]

Research of host defense peptides (HDPs) produced by helminths has been mostly neglected, although they are a segment of important cues within the tripartite communication among the parasite, host, and associated microbiota. Helminthic HDPs have low or no cyto- and genotoxicity toward host cells, in contrast to HDPs of free-living metazoans, which significantly hinders the use of the latter in preclinical and clinical studies in humans. Flatworm HDPs express a potent immunomodulatory activity, in contrast to nematode HDPs that express a potent antimicrobial activity, which might be a result of particular evolutionary adaptation, but this observation requires further investigation over a broader range of taxa. In silico prospecting of helminth genomes is a key for identifying known and novel HDP families, but it should be seconded by a functional screening approach. Parasitic helminths are destined to share niches with a variety of microbiota that inevitably influence their interaction with the host. To modulate the microbiome for their benefit and defend against pathogenic isolates, helminths have developed host defense peptides (HDPs) and proteins as integral elements of their immunity. These often exert a relatively nonspecific membranolytic activity toward bacteria, sometimes with limited or no toxicity toward host cells. With a few exceptions, such as nematode cecropin-like peptides and antibacterial factors (ABFs), helminthic HDPs are largely underexplored. This review scrutinizes current knowledge on the repertoire of such peptides in helminths and promotes their research as potential leads for an anti-infective solution to the burgeoning problem of antibiotic resistance. [ABSTRACT FROM AUTHOR]

Divergent evolution for more than 310 million years has resulted in an avian immune system that is complex and more compact than that of primates, sharing much of its structure and functions. Not surprisingly, well conserved ancient host defense molecules, such as defensins and cathelicidins, have diversified over time. In this review, we describe how evolution influenced the host defense peptides repertoire, its distribution, and the relationship between structure and biological functions. Marked features of primate and avian HDPs are linked to species-specific characteristics, biological requirements, and environmental challenge. • Genome size restraints governed diversification of avian cathelicidin genes and fewer β-defensin genes. • In contrast to primates, the cathelicidin repertoire is more diverse, but restricted in its distribution. • The conservation of multiple avian cathelicidin genes among different clades infer adaptations to niche biological functions. • β-sheet conservation in primate and avian β-defensins supports a convergent evolution towards receptor-mediated functions. [ABSTRACT FROM AUTHOR]

During recombinant protein expression in Pichia pastoris under the alcohol oxidase promoter, the culture supernatant developed green and yellow pigments, complicating downstream processing. Currently, a chromatography-based purification approach is one of the most efficient strategies to eliminate these pigments, but this method is complicated and often the most expensive step during processing. In this study, we designed a sequential cross-flow filtration by the combination of a microfiltration membrane with a pore size of 0.2 micrometers, followed by an ultrafiltration membrane with a molecular weight cut-off of 10 and 2 kilodaltons *Da), respectively, and finished with diafiltration of the 10 and 2 kDa retentate fractions. The results show that the microfiltration membrane eliminated host cellimpurities and significantly reduced the green pigment, which was measured using the 1976 CIE LAB system, while the 10 and 2 kDa membranes allowed the yellow pigment to pass through. The diafiltration steps also significantly reduced the yellow pigment. The host cell proteins were removed by recirculating in the retentate fraction of the 10 kDa membrane, while the 4.1 kDa target peptide, the candidate cell-penetrating antimicrobial peptide, was recovered by recirculation in the retentate fraction of the 21

New challenges in our society are appearing following the emergence of antibiotic-resistant microorganisms and the concerns about the use of antibiotics. Furthermore, food spoilage due to microorganisms leads to food waste of around 1.3 billion tons each year. Bioactive peptides obtained from food proteins, released by enzymatic hydrolysis or fermentation, can be employed as a natural alternative to antibiotics and as preservative agents, due to their ability to inhibit the growth of several microorganisms. Plant protein use in the food industry is increasing over animal protein, due to the environmental and animal welfare concerns of the population. Considering the concerns about the use antibiotics, their exploitation as a source of antimicrobial peptides is gaining interest in recent years. This review aims to summarize the published literature on protein hydrolysates as a source of antimicrobial peptides, obtained by enzymatic hydrolysis or fermentation of plants, their mechanisms, and the use in the food industry by producing fortified products or edible coatings. Antibacterial and antifungal plant peptides obtained by enzymatic hydrolysis are studied to a larger extent that antiviral peptides but further research concerning identification and applicability must be done. The results published show promising uses in the food industry in many areas such as fortification of foods or active packaging with dual bioactivity, related to the antimicrobial activity – avoiding food spoilage and antimicrobial resistance. [Display omitted] • Several plant antimicrobial peptides have been identified, mostly cationic. • Research on antifungal and antiviral plant peptides is scarce. • Analyses in real food models are required to claim the bioactivity and applicability. • Plant peptides offer a sustainable alternative to antibiotics and preservatives. [ABSTRACT FROM AUTHOR]

Dysfunction of the intestinal mucosal immune system and dysbiosis of the intestinal microflora can induce inflammatory bowel disease. However, drug-mediated clinical treatment remains a challenge due to its poor therapeutic efficacy and severe side effects. Herein, a ROS scavenging and inflammation-directed nanomedicine is designed and fabricated by coupling polydopamine nanoparticles with mCRAMP, an antimicrobial peptide, while wrapping macrophage membrane in the outer layer. The designed nanomedicine reduced the secretion of pro-inflammatory cytokines and elevate the expression of anti-inflammatory cytokine in vivo and in vitro inflammation models, demonstrating its significant ability of improving inflammatory responses. Importantly, the macrophage membrane encapsulated nanoparticles exhibit the obviously enhanced targeting performance in local inflamed tissues. Furthermore, the 16S rRNA sequencing of fecal microorganisms showed that probiotics increased and pathogenic bacteria were inhibited after oral delivery the nanomedicine, indicating that the designed nano platform played a significant role in optimizing intestinal microbiome. Taken together, the designed nanomedicine are not only easy to prepare and exhibit high biocompatibility, but also show the inflammatory targeting property, anti-inflammatory function and positive regulation of intestinal flora, thus providing a new idea for the intervention and treatment of colitis. Inflammatory bowel disease (IBD), a chronic and intractable disease, may lead to colon cancer in severe cases without effective treatment. However, clinical drugs are largely ineffective owing to insufficient therapeutic efficacies and side effects. Herein, we constructed a biomimetic polydopamine nanoparticle for oral administration to treat the IBD by modulating mucosal immune homeostasis and optimizing intestinal microorganisms. In vitro and in vivo experiments showed that the designed nanomedicine not only exhibits the anti-inflammatory function and inflammatory targeting property but also positively regulate the gut microflora. Taken together, the designed nanomedicine combined immunoregulation and intestinal microecology modulation to significantly enhance the therapeutic effect on colitis in mice, thus providing a new approach for the clinical treatment of colitis. [Display omitted] [ABSTRACT FROM AUTHOR]

Biofilms pose a great challenge for wound management. Herein, this study describes a near-infrared (NIR) light-responsive microneedle patch for on-demand release of antimicrobial peptide for treatment of wound biofilms. IR780 iodide as a photothermal conversion agent and molecularly engineered peptide W379 as an antimicrobial agent are loaded in dissolvable poly(vinylpyrrolidone) (PVP) microneedle patches followed by coating with a phase change material 1-tetradecanol (TD). After placing in an aqueous solution or biofilm containing wounds ex vivo and in vivo , upon exposure to NIR light, the incorporated IR780 induces light-to-heat conversion, causing the melting of TD. This leads to the dissolution of PVP microneedles, enabling the release of loaded W379 peptide from the microneedles into surrounding regions (e.g. , solution, biofilm, wound bed). Compared with traditional microneedle patches, NIR light responsive microneedle patches can program the release of antimicrobial peptide and show high antibacterial efficacy in vitro. Meanwhile, this work indicates that NIR light responsive TD-coated, W379-loaded PVP microneedle patches show excellent antibiofilm activities ex vivo and in vivo. Additionally, this microneedle system could be a promising platform for delivering other antimicrobial agents. This work demonstrates the fabrication of TD-coated, W379 and IR780 co-loaded PVP microneedle patches through the molding and spray coating. These microneedle patches display an excellent photothermal responsive antibacterial effect without significant cytotoxicity in vitro. In addition, these patches show high efficacy in combating wound biofilms ex vivo and in vivo. [Display omitted] [ABSTRACT FROM AUTHOR]

Cationic and amphiphilic peptides can be used as homing devices to accumulate conjugated antibiotics to bacteria-enriched sites and promote efficient microbial killing. However, just as important as tackling bacterial infections, is themodulation of the immune response in this complex microenvironment. In the present report, we designed a peptide chimaera called Chim2, formed by a membrane-active module, an enzyme hydrolysis site and a formyl peptide receptor 2 (FPR2) agonist. Thismoleculewas designed to adsorb onto bacterialmembranes, promote their lysis, and upon hydrolysis by local enzymes, release the FPR2 agonist sequence for activation and recruitment of immune cells. We synthesized the isolated peptide modules of Chim2 and characterized their biological activities independently and as a single polypeptide chain. We conducted antimicrobial assays, alongwith other tests aiming at the analyses of the cellular and immunological responses. In addition, assays using vesicles as models of eukaryotic and prokaryotic membranes were conducted and solution structures of Chim2 were generated by 1HNMR.Chim2 is antimicrobial, adsorbs preferentially to negatively charged vesicles while adopting an a-helix structure and exposes its disorganized tail to the solvent, which facilitates hydrolysis by tryptase-like enzymes, allowing the release of the FPR2 agonist fragment. This fragmentwas shown to induce accumulationof the cellular activation marker, lipid bodies, in mouse macrophages and the release of immunomodulatory interleukins. In conclusion, these data demonstrate that peptides with antimicrobial and immunomodulatory activities can be considered for further development as drugs. [ABSTRACT FROM AUTHOR]

In the current context of antibiotic resistance, the need to find alternative treatment strategies is urgent. Our research aimed to use synthetized aroylated phenylenediamines (APDs) to induce the expression of cathelicidin antimicrobial peptide gene (CAMP) to minimize the necessity of antibiotic use during infection. One of these compounds, HO53, showed promising results in inducing CAMP expression in bronchial epithelium cells (BCi-NS1.1 hereafter BCi). Thus, to decipher the cellular effects of HO53 on BCi cells, we performed RNA sequencing (RNAseq) analysis after 4, 8 and 24 h treatment of HO53. The number of differentially expressed transcripts pointed out an epigenetic modulation. Yet, the chemical structure and in silico modeling indicated HO53 as a histone deacetylase (HDAC) inhibitor. When exposed to a histone acetyl transferase (HAT) inhibitor, BCi cells showed a decreased expression of CAMP. Inversely, when treated with a specific HDAC3 inhibitor (RGFP996), BCi cells showed an increased expression of CAMP , indicating acetylation status in cells as determinant for the induction of the expression of the gene CAMP expression. Interestingly, a combination treatment with both HO53 and HDAC3 inhibitor RGFP966 leads to a further increase of CAMP expression. Moreover, HDAC3 inhibition by RGFP966 leads to increased expression of STAT3 and HIF1A , both previously demonstrated to be involved in pathways regulating CAMP expression. Importantly, HIF1α is considered as a master regulator in metabolism. A significant number of genes of metabolic enzymes were detected in our RNAseq data with enhanced expression conveying a shift toward enhanced glycolysis. Overall, we are demonstrating that HO53 might have a translational value against infections in the future through a mechanism leading to innate immunity strengthening involving HDAC inhibition and shifting the cells towards an immunometabolism, which further favors innate immunity activation. [Display omitted] • Antimicrobial inducer Aroylated Phenylenediamine HO53 also acts as HDAC inhibitor. • Specific HDAC3 inhibition further enhances HO53-induced CAMP expression. • HO53 increases expression of CAMP and metabolic genes, altering immunometabolism. • HO53 affects HDAC3/STAT3/HIF1α/ CAMP pathway. [ABSTRACT FROM AUTHOR]

Alcohol use among older adults is on the rise. This increase is clinically relevant as older adults are at risk for increased morbidity and mortality from many alcohol-related chronic diseases compared to younger patients. However, little is known regarding the synergistic effects of alcohol and age. There are intriguing data suggesting that aging may lead to impaired intestinal barrier integrity and dysbiosis of the intestinal microbiome, which could increase susceptibility to alcohol's negative effects. To study the effects of alcohol in age we exposed aged and young mice to 3 days of moderate ethanol and evaluated changes in gut parameters. We found that these levels of drinking do not have obvious effects in young mice but cause significant alcohol-induced gut barrier dysfunction and expression of the pro-inflammatory cytokine TNFα in aged mice. Ethanol-induced downregulation of expression of the gut-protective antimicrobial peptides Defa-rs1 , Reg3b , and Reg3g was observed in aged, but not young mice. Analysis of the fecal microbiome revealed age-associated shifts in microbial taxa, which correlated with intestinal and hepatic inflammatory gene expression. Taken together, these data demonstrate that age drives microbiome dysbiosis, while ethanol exposure in aged mice induces changes in the expression of antimicrobial genes important for separating these potentially damaging microbes from the intestinal lumen. These changes highlight potential mechanistic targets for prevention of the age-related exacerbation of effects of ethanol on the gut. • Aged mice display increased sensitivity to the gastrointestinal effects of ethanol. • Three days of ethanol binge increase intestinal barrier dysfunction and TNFα expression in aged, but not young mice. • Ethanol exposure in aged mice induces downregulation of intestinal antimicrobial peptides. • Age-specific changes in the fecal microbiome correlate with altered expression of cytokines and antimicrobial peptides. [ABSTRACT FROM AUTHOR]

Interactions between hyaluronan and the antimicrobial peptide cecropin B were studied in water and PBS using high-resolution ultrasonic spectroscopy and isothermal titration calorimetry. Although each technique is fundamentally different, they both gave identical results. It was found that the molecular weight of hyaluronan plays an important role in the interactions – in particular, the transition between the rod conformation and the random coil conformation. In water, interactions were saturated in a molar charge ratio of 1.5 and not 1.0 as expected. The later saturation of the interaction probably occurred either for steric reasons or due to the interaction between functional groups in the cecropin structure, which allowed complete dissociation of the antimicrobial peptide. In PBS, in contrast to water, no interactions were observed, irrespective of the molecular weight of hyaluronan. Thus, at a sufficiently high ionic strength, the interactions were suppressed. [Display omitted] • Hyaluronan interacts with cecropin B in water. • Interaction is suppressed in the physiological environment. • Interaction is significantly affected by the conformation of hyaluronan. • Saturation of interactions occurred after predicted charge equality. [ABSTRACT FROM AUTHOR]

Neutrophil extracellular traps (NETs) are structures consisting of decondensed chromatin with associated proteins, including histones and antimicrobial peptides, released from activated neutrophils. They are believed to be one of the body's first lines of defense against infectious agents. Despite their beneficial effect on the immune response process, some studies indicate that their excessive formation and the associated accumulation of extracellular DNA (eDNA) together with other polyelectrolytes (F-actin) plays an important role in the pathogenesis of many diseases. Thus NETs formation and removal are clinically significant. The monoclonal antibody 2C5 has strong specificity for intact nucleohistones (NS) and targets NS in NETs as we previously confirmed. Creation of a nano preparation that can specifically recognize and destroy NETs represents the aim for treatment many diseases. 2C5 antibody functionalized micelles coated with DNase I were created to achieve this aim. Schematic illustration of the 2C5 targeted MDM (mixed DNase I micelles) components (created by http://Biorender.com). [Display omitted] [ABSTRACT FROM AUTHOR]

Infections induced by intracellular pathogens are difficult to eradicate due to poor penetration of antimicrobials into cell membranes. It is of great importance to develop a new generation of antibacterial agents with dual functions of efficient cell penetration and bacterial inhibition. In this study, the association between hydrophobicity and cell-penetrating peptide delivery efficiency was investigated by fragment interception and hydrophobicity modification of natural porcine antimicrobial peptide PR-39 and the combination of cationic cell-penetrating peptide (R6) with antimicrobial peptide fragments modified with hydrophobic residues. The chimeric peptides P3I7 and P3L7, obtained through biofunctional screening, exhibited potent broad-spectrum antibacterial activity and low cytotoxicity. Moreover, P3I7 and P3L7 can effectively penetrate cells to eliminate intracellular pathogens mainly through endocytosis. The membrane destruction mechanism makes the peptides fast sterilizers and less prone to developing drug resistance. Finally, their good biocompatibility and antibacterial infection effects were verified in mice and piglets. To conclude, the chimeric peptides P3I7 and P3L7 show great potential as affordable and effective antimicrobial agents and may serve as ideal candidates for the treatment of intracellular bacterial infections. The low permeability of antibacterial drugs makes infections induced by intracellular bacteria extremely difficult to treat. To address this issue, we designed chimeric peptides with dual cell-penetrating and antibacterial functions. The active peptides P3I7 and P3L7, acquired through functional screening have strong broad-spectrum antibacterial activity and powerful bactericidal effects against intracellular Staphylococcus aureus. The membrane permeation mechanism of P3I7 and P3L7 against bacteria endows fast bactericidal activity with low drug resistance. The biosafety and antibacterial activity of P3I7 and P3L7 were also validated by in vivo trials. This study provides an ideal drug candidate against intracellular bacterial infections. [Display omitted] [ABSTRACT FROM AUTHOR]

Tryptocidine C (TpcC), a Trp-rich cyclodecapeptide is a minor constituent in the antibiotic tyrothricin complex from Brevibacillus parabrevis. TpcC possesses a high tendency to oligomerise in aqueous solutions and dried TpcC forms distinct self-assembled nanoparticles. High-resolution scanning electron microscopy revealed the influence of different ethanol:water solvent systems on TpcC self-assembly, with the TpcC, dried from a high concentration in 15% ethanol, primarily assembling into small nanospheres with 24.3 nm diameter and 0.05 polydispersity. TpcC at 16 μM, near its CMC, formed a variety of structures such as small nanospheres, large dense nanospheroids and facetted 3-D-crystals, as well as sheets and coarse carpet-like structures which depended on ethanol concentration. Drying 16 μM TpcC from 75% ethanol resulted in highly facetted 3-D crystals, as well as small nanospheres, while those in 10% ethanol preparation had less defined facets. Drying from 20 to 50% ethanol led to polymorphic architectures with a few defined nanospheroids and various small nanoparticles, imbedded in carpet- and sheet-like structures. These polymorphic surface morphologies correlated with maintenance of fluorescence properties and the surface-derived antibacterial activity against Staphylococcus aureus over time, while there was a significant change in fluorescence and loss in activity in the 10% and 75% preparations where 3-D crystals were observed. This indicated that TpcC oligomerisation in solutions with 20–50% ethanol leads to metastable structures with a high propensity for release of antimicrobial moieties, while those leading to crystallisation limit active moieties release. TpcC nano-assemblies can find application in antimicrobial coatings, surface disinfectants, food packaging and wound healing materials. [Display omitted] • The type of self-assembly structures of the Trp-rich cyclodecapeptide, TpcC can be manipulated via concentration, time in solution and [ethanol]. • TpcC assemble into sheets and small nanospheres with 24 nm diameter which act the template and feedstock for facetted 3-D crystals. • The antimicrobial activity of surface deposited TpcC is influenced by the nanostructure character and its metastability. • This first study reveal the correlation between of TpcC solution phase oligomerisation, surface structures and its antimicrobial activity. [ABSTRACT FROM AUTHOR]