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Muga is a semi-domesticated multi-voltine silkworm cultivated in different seasons of the year. Crops are reared in specific months and are traditionally named Jarua (spring), Chotua, Bohogua, Jethua, Aherua, Saonia, Bhodia, Ahinia, Katia (autumn) and Aghonia. Seasonal variation forces silkworm larvae to face harsh environmental conditions manifested by generation of oxidative stress and occurrence of microbial diseases. Anti-oxidant enzymes like superoxide dismutase (SOD), catalase (CAT), and glutathione-S-transferase (GST) along with anti-microbial peptides like cecropin, gloverin, and defensin in lepidopterans play a significant role in fighting against free radicals as well as disease-causing microbes, and hence this study has focussed on the expression of anti-oxidant enzyme activity and different anti-microbial peptides in three distinct crops, namely, Jarua and Aherua (seed crops) and Katia (commercial crop) collected from the same site, Khanapara (26°09′34.62″N; 91°41′27.23″E) in Assam, India. The results of biochemical enzyme assays revealed the Jarua crop to possess the highest enzyme activity with respect to SOD and GST in particular, followed by the Katia and Aherua crops. Malondialdehyde lipid peroxidase and reduced glutathione were also expressed notably in the Jarua crop. The quantitative polymerase chain reaction (qPCR) study involving analysis of anti-microbial peptide (AMP) expression revealed the Jarua crop to possess the highest expression of three AMPs, viz., gloverin, moricin 1 and gallerimycin pro, followed by Katia with the highest expression of attacin and defensin. Total haemocyte count revealed Jarua to induce the least haemocyte count, Katia to induce moderate, and Aherua to induce the highest count. Thus, our findings indicate that although the Jarua crop is considered a seed crop, it can be reconsidered as a commercial crop due to its high concentration of anti-oxidant enzymes and higher expression of AMPs. This study was carried out to focus on the immunological response of the muga silkworm with varying seasons. Their innate immunity helps them fight against diseases that emerge under different environmental conditions in which anti-microbial peptides play a crucial role. Muga larvae are reared for silk and are used commercially to produce various commodities for which the best silk thread should always be identified to obtain a higher market value. [ABSTRACT FROM AUTHOR]

Since infections with antibiotic-resistant bacteria cause increasing problems worldwide, the identification of alternative therapies is of great importance. Plantderived bioactives, including allyl-isothiocyanate (AITC), have received attention for their antimicrobial properties. The present study therefore investigates the impact of AITC on survival and antimicrobial peptide (AMP) levels in Drosophila melanogaster challenged with the fly pathogenic bacteria Pectobacterium carotovorum subsp. carotovorum and Leuconostoc pseudomesenteroides. AITC, a sulfur-containing compound derived from glucosinolates, exhibits antimicrobial properties and has been suggested to modulate AMP expression. By using D. melanogaster, we demonstrate that AITC treatment resulted in a concentration-dependent decrease of survival rates among female flies, particularly in the presence of the Gramnegative bacterium Pectobacterium carotovorum subsp. carotovorum, whereas AITC did not affect survival in male flies. Despite the ability of isothiocyanates to induce AMP expression in cell culture, we did not detect significant changes in AMP mRNA levels in infected flies exposed to AITC. Our findings suggest sex-specific differences in response to AITC treatment and bacterial infections, underlining the complexity of host--pathogen interactions and potential limitations of AITC as a preventive or therapeutic compound at least in D. melanogaster models of bacterial infections. [ABSTRACT FROM AUTHOR]

Introduction: Aging studies in humans and mice have played a key role in understanding the intestinal microbiome and an increased abundance of “inflammaging” Gram-negative (Gn) bacteria. The mechanisms underlying this inflammatory profile in the aging microbiome are unknown. We tested the hypothesis that an aging-related decrease in colonic crypt epithelial cell anti)microbial peptide (AMP) gene expression could promote colonic microbiome inflammatory Gn dysbiosis and inflammaging. Methods: As a model of aging, C57BL/6J mice fecal (colonic) microbiota (16S) and isolated colonic crypt epithelial cell gene expression (RNA-seq) were assessed at 2 months (mth) (human: 18 years old; yo), 15 mth (human: 50 yo), and 25 mth (human: 84 yo). Informatics examined aging-related microbial compositions, differential colonic crypt epithelial cell gene expressions, and correlations between colonic bacteria and colonic crypt epithelial cell gene expressions. Results: Fecal microbiota exhibited significantly increased relative abundances of pro-inflammatory Gn bacteria with aging. Colonic crypt epithelial cell gene expression analysis showed significant age-related downregulation of key AMP genes that repress the growth of Gn bacteria. The aging-related decrease in AMP gene expressions is significantly correlated with an increased abundance in Gn bacteria (dysbiosis), loss of colonic barrier gene expression, and senescence- and inflammation-related gene expression. Conclusion: This study supports the proposed model that aging-related loss of colonic crypt epithelial cell AMP gene expression promotes increased relative abundances of Gn inflammaging-associated bacteria and gene expression markers of colonic inflammaging. These data may support new targets for aging-related therapies based on intestinal genes and microbiomes. [ABSTRACT FROM AUTHOR]

Since infections with antibiotic-resistant bacteria cause increasing problems worldwide, the identification of alternative therapies is of great importance. Plant-derived bioactives, including allyl-isothiocyanate (AITC), have received attention for their antimicrobial properties. The present study therefore investigates the impact of AITC on survival and antimicrobial peptide (AMP) levels in Drosophila melanogaster challenged with the fly pathogenic bacteria Pectobacterium carotovorum subsp. carotovorum and Leuconostoc pseudomesenteroides. AITC, a sulfur-containing compound derived from glucosinolates, exhibits antimicrobial properties and has been suggested to modulate AMP expression. By using D. melanogaster, we demonstrate that AITC treatment resulted in a concentration-dependent decrease of survival rates among female flies, particularly in the presence of the Gram-negative bacterium Pectobacterium carotovorum subsp. carotovorum, whereas AITC did not affect survival in male flies. Despite the ability of isothiocyanates to induce AMP expression in cell culture, we did not detect significant changes in AMP mRNA levels in infected flies exposed to AITC. Our findings suggest sex-specific differences in response to AITC treatment and bacterial infections, underlining the complexity of host–pathogen interactions and potential limitations of AITC as a preventive or therapeutic compound at least in D. melanogaster models of bacterial infections.

The incidence of oro-dental disorders has emerged as a serious threat to the healthcare sector owing to the increasing complexity of the oral microbiome. Significant advances in biomaterial research have led to the advent of a plethora of drug delivery systems including nanocarriers, dendrimers, hydrogels and other kinds of stimuli-responsive polymeric biomaterials. Anti-microbial peptides (AMPs) have engendered considerable interest in the past decades as potential alternatives to traditional disinfecting agents and also emerged as potent antibiofilm agents. Among the most viable approaches in targeted drug delivery, hydrogels incorporated with AMPs are emerging as bio-functional platforms yielding increased stability and bioavailability. The antibacterial and antibiofilm activities of Nisin are studied using microbiological methods followed by the synthesis of Nisin loaded PVA-Alginate hydrogel for dental treatment. The physicochemical characteristics of Nisin loaded hydrogel were done by swelling behavior, release kinetics assay, FTIR spectroscopic methods and cytotoxicity studies. Nisin showed antibacterial activity towards clinical isolates of drug-resistant bacteria and the antibiofilm and anti-adhesion studies demonstrated that Nisin could control the bacterial count in the test sample. The polymerization of Nisin into hydrogels was done and the physico-chemical characterization of Nisin loaded hydrogel network could be envisaged as a potential drug delivery platform for oral infections. Nisin loaded PVA-Alginate biocompatible hydrogel exhibited apparent swellable, flexible, nonhaemolytic materials and active antimicrobial and nontoxic materials. Physicochemical properties of these Nisin loaded PVA-Alginate biocompatible hydrogels have great potential in biomaterial-based drug delivery systems in controlling the growth and proliferation of major oro-dental pathogens. This could be exploited for the temporary biocompatible dental filling materials to treat the caries. Exploring potential nisin loaded hydrogel delivery systems will provide a brighter future of more friendly, effective and personalized treatment to deal with dental caries.

Introduction: Aging studies in humans and mice have played a key role in understanding the intestinal microbiome and an increased abundance of “inflammaging” Gram-negative (Gn) bacteria. The mechanisms underlying this inflammatory profile in the aging microbiome are unknown. We tested the hypothesis that an aging-related decrease in colonic crypt epithelial cell anti-microbial peptide (AMP) gene expression could promote colonic microbiome inflammatory Gn dysbiosis and inflammaging.Methods: As a model of aging, C57BL/6J mice fecal (colonic) microbiota (16S) and isolated colonic crypt epithelial cell gene expression (RNA-seq) were assessed at 2 months (mth) (human: 18 years old; yo), 15 mth (human: 50 yo), and 25 mth (human: 84 yo). Informatics examined aging-related microbial compositions, differential colonic crypt epithelial cell gene expressions, and correlations between colonic bacteria and colonic crypt epithelial cell gene expressions.Results: Fecal microbiota exhibited significantly increased relative abundances of pro-inflammatory Gn bacteria with aging. Colonic crypt epithelial cell gene expression analysis showed significant age-related downregulation of key AMP genes that repress the growth of Gn bacteria. The aging-related decrease in AMP gene expressions is significantly correlated with an increased abundance in Gn bacteria (dysbiosis), loss of colonic barrier gene expression, and senescence- and inflammation-related gene expression.Conclusion: This study supports the proposed model that aging-related loss of colonic crypt epithelial cell AMP gene expression promotes increased relative abundances of Gn inflammaging-associated bacteria and gene expression markers of colonic inflammaging. These data may support new targets for aging-related therapies based on intestinal genes and microbiomes.

The incidence of oro-dental disorders has emerged as a serious threat to the healthcare sector owing to the increasing complexity of the oral microbiome. Significant advances in biomaterial research have led to the advent of a plethora of drug delivery systems including nanocarriers, dendrimers, hydrogels and other kinds of stimuli-responsive polymeric biomaterials. Anti-microbial peptides (AMPs) have engendered considerable interest in the past decades as potential alternatives to traditional disinfecting agents and also emerged as potent antibiofilm agents. Among the most viable approaches in targeted drug delivery, hydrogels incorporated with AMPs are emerging as bio-functional platforms yielding increased stability and bioavailability. The antibacterial and antibiofilm activities of Nisin are studied using microbiological methods followed by the synthesis of Nisin loaded PVA-Alginate hydrogel for dental treatment. The physico-chemical characteristics of Nisin loaded hydrogel were done by swelling behavior, release kinetics assay, FTIR spectroscopic methods and cytotoxicity studies. Nisin showed antibacterial activity towards clinical isolates of drug-resistant bacteria and the antibiofilm and anti-adhesion studies demonstrated that Nisin could control the bacterial count in the test sample. The polymerization of Nisin into hydrogels was done and the physico- chemical characterization of Nisin loaded hydrogel network could be envisaged as a potential drug delivery platform for oral infections. Nisin loaded PVA-Alginate biocompatible hydrogel exhibited apparent swellable, flexible, nonhaemolytic materials and active antimicrobial and nontoxic materials. Physicochemical properties of these Nisin loaded PVA-Alginate biocompatible hydrogels have great potential in biomaterial-based drug delivery systems in controlling the growth and proliferation of major oro-dental pathogens. This could be exploited for the temporary biocompatible dental filling materials to treat the caries. Exploring potential nisin loaded hydrogel delivery systems will provide a brighter future of more friendly, effective and personalized treatment to deal with dental caries. [ABSTRACT FROM AUTHOR]

BackgroundThe DNA-binding peptide LL37 is a suspected autoantigen in psoriasis. It can be found in neutrophil extracellular traps (NETs) which have been suggested to play a role in the pathogenesis of the disease. Citrullination, the conversion of peptidyl-arginine into peptidyl-citrulline, can be implicated in the formation of NETs. We hypothesized that citrullination increases LL37 immunogenicity and that NETs are a source of LL37.ObjectivesWe aimed to characterize cytokine responses of B cells and T cells to native and citrullinated LL37 (citLL37) and determine the prevalence and composition of circulating NETs in patients with psoriasis and healthy blood donors (HDs).MethodsMononuclear cells (MNCs) and serum were isolated from 20 HDs and 20 patients with psoriasis. The MNCs were stimulated with native LL37 and citLL37 and the proportion of cytokine-positive B cells and T cells was determined by flow cytometry. Circulating antibodies against native LL37 and citLL37 as well as circulating NETs were measured by ELISA, as was the content of LL37, citLL37, and IgG in the NETs.ResultsCitLL37, but not native LL37, induced IFN-γ-production by T cells and B cells from psoriasis patients, as well as IL-10-production by the patients’ CD4+ T cells. Serum from 40% of patients and 55% of HDs contained circulating NETs, of which 63% and 27%, respectively, contained LL37. Only two patients had NETs containing citLL37 and IgG antibodies were found in NETs from three patients and one HD. Post-hoc analysis of the cytokines produced by B cells and T cells after stimulation with citLL37 revealed two clusters of patients consisting of 10 high-responders and 9 low-responders. The high-responders were those that had circulating NETs in combination with an earlier age of onset of the disease.ConclusionCitrullinated but not native LL37 elicits IFN-γ-responses by T cells and B cells from psoriasis patients, particularly those with circulating NETs and early disease onset, suggesting a role of citLL37 as an autoantigen in this subgroup of patients.

Summary: Background: In preclinical models of Type 1 Diabetes (T1D) the integrity of the gut barrier (GB) is instrumental to avoid dysregulated crosstalk between the commensal microbiota and immune cells and to prevent autoimmunity. The GB is composed of the intestinal epithelial barrier (IEB) and of the mucus layer containing mucins and antimicrobial peptides (AMPs) that are crucial to maintain immune tolerance. In preclinical models of T1D the alterations of the GB primarily affect the mucus layer. In human T1D increased gut permeability and IEB damage have been demonstrated but the integrity of the mucus layer was never assessed. Methods: We evaluated GB integrity by measuring serological markers of IEB damage (serological levels of zonulin) and bacterial translocation such as lipopolysaccharide binding protein (LBP) and myeloid differentiation protein 2 (MD2), and mRNA expression of tight junction proteins, mucins and AMPs in intestinal tissue of T1D patients and healthy controls (HC). Simultaneously, we performed immunological profiling on intestinal tissue and 16S rRNA analysis on the mucus-associated gut microbiota (MAGM). Findings: Our data show a GB damage with mucus layer alterations and reduced mRNA expression of several mucins (MUC2, MUC12, MUC13, MUC15, MUC20, MUC21) and AMPs (HD4 and HD5) in T1D patients. Mucus layer alterations correlated with reduced relative abundance of short chain fatty acids (SCFA)-producing bacteria such as Bifidobacterium dentium, Clostridium butyricum and Roseburia intestinalis that regulate mucin expression and intestinal immune homeostasis. In T1D patients we also found intestinal immune dysregulation with higher percentages of effector T cells such as T helper (Th) 1, Th17 and TNF-α+ T cells. Interpretation: Our data show that mucus layer alterations are present in T1D subjects and associated with dysbiosis and immune dysregulation. Funding: Research Grants from the Juvenile Diabetes Foundation (Grant 1-INO-2018-640-A-N to MF and 2-SRA-2019-680-S-B to JD) and from the Italian Ministry of Health (Grant RF19-12370721 to MF).

Antibiotic resistance is rapidly increasing, and new anti-infective therapies are urgently needed. In this regard, antimicrobial peptides (AMPs) may represent potential candidates for the treatment of infections caused by multiresistant microorganisms. In this narrative review, we reported the experience of our research group over 20 years. We described the AMPs we evaluated against Gram-positive, Gram-negative, and fungi. In conclusion, our experience shows that AMPs can be a key option for treating multiresistant infections and overcoming resistance mechanisms. The combination of AMPs allows antibiotics and antifungals that are no longer effective to exploit the synergistic effect by restoring their efficacy. A current limitation includes poor data on human patients, the cost of some AMPs, and their safety, which is why studies on humans are needed as soon as possible. [ABSTRACT FROM AUTHOR]

Laura I Ortiz-Lopez,1,2 Vivek Choudhary,1,3 Wendy B Bollag1,3– 5 1Department of Physiology, Medical College of Georgia at Augusta University, Augusta, GA, 30912, USA; 2Department of Neuroscience and Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA, 30912, USA; 3Charlie Norwood VA Medical Center, Augusta, GA, 30904, USA; 4Department of Dermatology, Medical College of Georgia at Augusta University, Augusta, GA, 30912, USA; 5Department of Medicine, Medical College of Georgia at Augusta University, Augusta, GA, 30912, USACorrespondence: Wendy B Bollag, Department of Physiology, Medical College of Georgia at Augusta University, 1120 15th Street, Augusta, GA, 30912, USA, Tel +1 706 721-0698, Fax +1 706 721-7299, Email wbollag@augusta.eduAbstract: Psoriasis is a complex disease triggered by genetic, immunologic, and environmental stimuli. Many genes have been linked to psoriasis, like the psoriasis susceptibility genes, some of which are critical in keratinocyte biology and epidermal barrier function. Still, the exact pathogenesis of psoriasis is unknown. In the disease, the balance between the proliferative and differentiative processes of keratinocytes becomes altered. Multiple studies have highlighted the role of dysregulated immune cells in provoking the inflammatory responses seen in psoriasis. In addition to immune cells, accumulating evidence shows that keratinocytes are involved in psoriasis pathogenesis, as discussed in this review. Although certain immune cell-derived factors stimulate keratinocyte hyperproliferation, activated keratinocytes can also produce anti-microbial peptides, cytokines, and chemokines that can promote their proliferation, as well as recruit immune cells to help initiate and reinforce inflammatory feedback loops. Psoriatic keratinocytes also show intrinsic differences from normal keratinocytes even after removal from the in vivo inflammatory environment; thus, psoriatic keratinocytes have been found to exhibit abnormal calcium metabolism and possible epigenetic changes that contribute to psoriasis. The Koebner phenomenon, in which injury promotes the development of psoriatic lesions, also provides evidence for keratinocytes’ contributions to disease pathogenesis. Furthermore, transgenic mouse studies have confirmed the importance of keratinocytes in the etiology of psoriasis. Finally, in addition to immune cells and keratinocytes, data in the literature support roles for other cell types, tissues, and systems in psoriasis development. These other contributors are all potential targets for therapies, suggesting the importance of a holistic approach when treating psoriasis.Keywords: anti-microbial peptides, inflammation, innate immunity, Koebner phenomenon, skin

Acanthamoeba is a free-living protozoan known to cause keratitis most commonly, especially among contact lens wearers. Treatment of Acanthamoeba keratitis is challenging as Acanthamoeba can encyst from the active form, a trophozoite, into a hibernating cyst that is refractory to antibiotics and difficult to kill; therefore, there is a need for more effective anti-amoebic strategies. In this study, we have evaluated the anti-amoebic activity of the antimicrobial peptide mimic RK-758 against Acanthamoeba castellanii. RK-758 peptidomimetic was subjected to biological assays to investigate its amoebicidal, amoebistatic, anti-encystation, and anti-excystation effects on A. castellanii. The anti-amoebic activity of the peptide mimic RK-758 was compared with chlorhexidine against the Acanthamoeba castellanii ATCC30868 and Acanthamoeba castellanii 044 (a clinical strain) with the concentrations of both ranging from 125 µM down to 7.81 µM. All experiments were performed in duplicate with three independent replicates. The data were represented as mean ± SE and analysed using a two-sample t-test and two-tailed distributions. A p < 0.05 was considered statistically significant. The peptidomimetic RK-758 had anti-Acanthamoeba activity against both trophozoites and cysts in a dose-dependent manner. The RK-758 had amoebicidal and growth inhibitory activities of ≥50% at a concentration between 125 µM and 15.6 µM against the trophozoites of both Acanthamoeba strains. Inhibitory effects on the cyst formation and trophozoite re-emergence from cysts were noted at similar concentrations. Chlorhexidine had 50% activity at 7.81 µM and above against the trophozoites and cysts of both strains. In the haemolysis assay, the RK-758 lysed horse RBCs at concentrations greater than 50 µM whereas lysis occurred at concentrations greater than 125 µM for the chlorhexidine. The peptidomimetic RK-758, therefore, has activity against both the trophozoite and cyst forms of Acanthamoeba and has the potential to be further developed as an anti-microbial agent against Acanthamoeba. RK-758 may also have use as an anti-amoebic disinfectant in contact lens solutions. [ABSTRACT FROM AUTHOR]

Abstract Background Although Escherichia coli has been widely used for the expression of exogenous proteins, the secretory expression in this system is still a big obstacle. As one of the most important secretion pathways, hemolysin A (HlyA) system of E. coli can transport substrates directly from the cytoplasm to extracellular medium without the formation of any periplasmic intermediate, making it an ideal candidate for the development of the secretory production platform for exogenous proteins. Results In this work, we developed a novel production platform, THHly, based on the HlyA secretion system, and explored its applications in the efficient preparation and quick detection of tag peptides and anti-microbial peptides. In this novel platform the signal sequence of HlyA is fused to the C-terminal of target peptide, with Tobacco Etch Virus (TEV) protease cleavage site and 6*His tag between them. Five tag peptides displayed good secretory properties in E. coli BL21 (DE3), among which T7 tag and S tag were obtained by two rounds of purification steps and TEV cleavage, and maintained their intrinsic immunogenicity. Furthermore, Cecropin A and Melittin, two different types of widely explored anti-microbial peptides, were produced likewise and verified to possess anti-microbial/anti-tumor bioactivities. No significant bacterial growth inhibition was observed during the fusion protein expression, indicating that the fusion form not only mediated the secretion but also decreased the toxicity of anti-microbial peptides (AMPs) to the host bacteria. To the best of our knowledge, this is the first report to achieve the secretory expression of these two AMPs in E. coli with considerable potential for manufacturing and industrialization purposes. Conclusions The results demonstrate that the HlyA based novel production platform of E. coli allowed the efficient secretory production and purification of peptides, thus suggesting a promising strategy for the industrialized production of peptide pharmaceuticals or reagents. Graphical Abstract

Periodontal diseases are common inflammatory conditions characterized by bone loss in response to simultaneous bacterial aggression and host defenses. The etiology of such diseases is still not completely understood, however. It has been shown that specific pathogens involved in the build-up of dysbiotic biofilms participate actively in the establishment of periodontitis. This multifactorial pathology also depends on environmental factors and host characteristics, especially defenses. The immune response to the pathogens seems to be critical in preventing the disease from starting but also contributes to tissue damage. It is known that small molecules known as antimicrobial peptides (AMPs) are key actors in the innate immune response. They not only target microbes, but also act as immuno-modulators. They can help to recruit or activate cells such as neutrophils, monocytes, dendritic cells, or lymphocytes. AMPs have already been described in the periodontium, and their expression seems to be connected to disease activity. Alpha and beta defensins and LL37 are the AMPs most frequently linked to periodontitis. Additionally, leukocyte infiltrates, especially B-cells, have also been linked to the severity of periodontitis. Indeed, the particular subpopulations of B-cells in these infiltrates have been linked to inflammation and bone resorption. A link between B-cells and AMP could be relevant to understanding B-cells' action. Some AMP receptors, such as chemokines receptors, toll-like receptors, or purinergic receptors, have been shown to be expressed by B-cells. Consequently, the action of AMPs on B--cell subpopulations could participate to B-cell recruitment, their differentiation, and their implication in both periodontal defense and destruction. [ABSTRACT FROM AUTHOR]

The rise of antibiotic resistant bacteria due to overuse and misuse of antibiotics in medicine and dentistry is a growing concern. New approaches are needed to combat antibiotic resistant (AR) bacterial infections. There are a number of methods available and in development to address AR infections. Dentists conventionally use chemicals such as chlorohexidine and calcium hydroxide to kill oral bacteria, with many groups recently developing more biocompatible antimicrobial peptides (AMPs) for use in the oral cavity. AMPs are promising candidates in the treatment of (oral) infections. Also known as host defense peptides, AMPs have been isolated from animals across all kingdoms of life and play an integral role in the innate immunity of both prokaryotic and eukaryotic organisms by responding to pathogens. Despite progress over the last four decades, there are only a few AMPs approved for clinical use. This review summarizes an Introduction to Oral Microbiome and Oral Infections, Traditional Antibiotics and Alternatives & Antimicrobial Peptides. There is a focus on cationic AMP characteristics and mechanisms of actions, and an overview of animal-derived natural and synthetic AMPs, as well as observed microbial resistance. [ABSTRACT FROM AUTHOR]

Diabetic hyperglycemia delays wound healing, leading to serious consequences. Topical antibiotics can reduce the risk of a wound infection during healing; nevertheless, the microbial fight against antibiotics brings about public health challenges. Anti-microbial peptides (AMPs) belong to a novel class of drug that is used to prevent and treat systemic and topical infections. The aim of the current work was to achieve better wound healing in diabetic rats by conjugating the anti-microbial peptide "apamin" (APA) with the broad-spectrum antibiotic "ceftriaxone" (CTX) to form a nanocomplex. The CTX–APA nanoconjugate formulation was optimized using a Box–Behnken design. The optimized CTX–APA nanoconjugate formulation was evaluated for its size and zeta potential, and was then examined using transmission electron microscopy (TEM). The CTX–APA nanoconjugate was loaded onto a hydroxypropyl methylcellulose (2% w/v)-based hydrogel. It was observed that the application of the CTX–APA nanocomplex on the wounded skin of diabetic rats accelerated the regeneration of the epithelium, granulation tissue formation, epidermal proliferation, and keratinization. The nanocomplex was capable of significantly reducing the expression of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6), while increasing the expression of transforming growth factor beta-1 (TGF-β1) as well as the angiogenic markers: hypoxia-inducible factor 1-alpha (HIF-1α) and vascular endothelial growth factor (VEGF). Conclusively, the application of an ion-paired CTX–APA nanocomplex enhances wound healing in diabetic rats. [ABSTRACT FROM AUTHOR]

Periodontal diseases are common inflammatory conditions characterized by bone loss in response to simultaneous bacterial aggression and host defenses. The etiology of such diseases is still not completely understood, however. It has been shown that specific pathogens involved in the build-up of dysbiotic biofilms participate actively in the establishment of periodontitis. This multifactorial pathology also depends on environmental factors and host characteristics, especially defenses. The immune response to the pathogens seems to be critical in preventing the disease from starting but also contributes to tissue damage. It is known that small molecules known as antimicrobial peptides (AMPs) are key actors in the innate immune response. They not only target microbes, but also act as immuno-modulators. They can help to recruit or activate cells such as neutrophils, monocytes, dendritic cells, or lymphocytes. AMPs have already been described in the periodontium, and their expression seems to be connected to disease activity. Alpha and beta defensins and LL37 are the AMPs most frequently linked to periodontitis. Additionally, leukocyte infiltrates, especially B-cells, have also been linked to the severity of periodontitis. Indeed, the particular subpopulations of B-cells in these infiltrates have been linked to inflammation and bone resorption. A link between B-cells and AMP could be relevant to understanding B-cells’ action. Some AMP receptors, such as chemokines receptors, toll-like receptors, or purinergic receptors, have been shown to be expressed by B-cells. Consequently, the action of AMPs on B—cell subpopulations could participate to B-cell recruitment, their differentiation, and their implication in both periodontal defense and destruction.

Filamentous fungi are a large and diverse taxonomically group of microorganisms found in all habitats worldwide. They grow as a network of cells called hyphae. Since filamentous fungi live in very diverse habitats, they produce different enzymes to degrade material for their living, for example hydrolytic enzymes to degrade various kinds of biomasses. Moreover, they produce defense proteins (antimicrobial peptides) and proteins for attaching surfaces (hydrophobins). Many of them are easy to cultivate in different known setups (submerged fermentation and solid-state fermentation) and their secretion of proteins and enzymes are often much larger than what is seen from yeast and bacteria. Therefore, filamentous fungi are in many industries the preferred production hosts of different proteins and enzymes. Edible fungi have traditionally been used as food, such as mushrooms or in fermented foods. New trends are to use edible fungi to produce myco-protein enriched foods. This review gives an overview of the different kinds of proteins, enzymes, and peptides produced by the most well-known fungi used as cell factories for different purposes and applications. Moreover, we describe some of the challenges that are important to consider when filamentous fungi are optimized as efficient cell factories. [ABSTRACT FROM AUTHOR]

Interleukin-6 (IL-6) is a multifunctional cytokine involved in innate and acquired immunity. In mammals, IL-6 can induce the secretion of acute-phase proteins (APPs) and antimicrobial peptides, activate macrophages and neutrophils, and regulate the expression of inflammatory cytokines in innate immunity, resulting in enhanced resistance to invading pathogens. In teleost fish, the functions of IL-6 are still unclear. In our previous studies using orange-spotted grouper (Epinephelus coioides) as an animal model, we demonstrated that IL-6 induces the Th2 pathway in acquired immunity and enhances antibody production. In the current study, we evaluated in greater detail the mechanisms underlying the functions of IL-6 in innate immunity. In gene expression assays, the expression of fish inflammatory cytokine genes (e.g., ifnγ, ccl4, il1β, tnfα 1, and tnfα 2), antimicrobial peptide genes (e.g., epinecidin 1, hepcidin 1, and hepcidin 2), and genes related to innate immunity (e.g., mx, crp, and complement C3) was higher in fish injected with recombinant IL-6 (rIL-6). The serum from those same fish also presented higher antibacterial activity and higher phagocytosis activity of leukocytes. In a pathogen challenge involving Vibrio campbellii and nervous necrosis virus, fish injected with rIL-6 presented a higher survival rate and lower pathogen count in the body. These results demonstrate that the administration of rIL-6 can activate innate immune responses and enhance resistance against invading bacterial or viral pathogens.

Bark protects the tree against environmental insults. Here, we analyzed whether this defensive strategy could be utilized to broadly enhance protection against colitis. As a proof of concept, we show that exosome‐like nanoparticles (MBELNs) derived from edible mulberry bark confer protection against colitis in a mouse model by promoting heat shock protein family A (Hsp70) member 8 (HSPA8)‐mediated activation of the AhR signaling pathway. Activation of this pathway in intestinal epithelial cells leads to the induction of COP9 Constitutive Photomorphogenic Homolog Subunit 8 (COPS8). Utilizing a gut epithelium‐specific knockout of COPS8, we demonstrate that COPS8 acts downstream of the AhR pathway and is required for the protective effect of MBELNs by inducing an array of anti‐microbial peptides. Our results indicate that MBELNs represent an undescribed mode of inter‐kingdom communication in the mammalian intestine through an AhR‐COPS8‐mediated anti‐inflammatory pathway. These data suggest that inflammatory pathways in a microbiota‐enriched intestinal environment are regulated by COPS8 and that edible plant‐derived ELNs may hold the potential as new agents for the prevention and treatment of gut‐related inflammatory disease. Synopsis: Mulberry bark derived exosome‐like nanoparticles (MBELNs) prevent gut inflammation via plant heat shock protein HSPA8‐mediated activation of AhR/COPS8 pathways. Treatment with MBELNs promotes the restoration of gut microbiome homeostasis, ameliorating intestinal inflammatory pathologies. Mulberry bark derived exosome‐like nanoparticles (MBELNs) prevent mouse colitis via the AhR/COPS8 pathway.Binding of MBELN‐derived heat shock protein HSPA8 to AhR leads to the activation of AhR signaling.Activation of AhR leads to the induction of an array of anti‐microbial peptides (AMPs) via COP9/COPS8.AhR/COPS8‐dependent induction of AMPs inhibits intestinal inflammation and alters fecal gut microbiota composition. [ABSTRACT FROM AUTHOR]

Background: Although Escherichia coli has been widely used for the expression of exogenous proteins, the secretory expression in this system is still a big obstacle. As one of the most important secretion pathways, hemolysin A (HlyA) system of E. coli can transport substrates directly from the cytoplasm to extracellular medium without the formation of any periplasmic intermediate, making it an ideal candidate for the development of the secretory production platform for exogenous proteins. Results: In this work, we developed a novel production platform, THHly, based on the HlyA secretion system, and explored its applications in the efficient preparation and quick detection of tag peptides and anti-microbial peptides. In this novel platform the signal sequence of HlyA is fused to the C-terminal of target peptide, with Tobacco Etch Virus (TEV) protease cleavage site and 6*His tag between them. Five tag peptides displayed good secretory properties in E. coli BL21 (DE3), among which T7 tag and S tag were obtained by two rounds of purification steps and TEV cleavage, and maintained their intrinsic immunogenicity. Furthermore, Cecropin A and Melittin, two different types of widely explored anti-microbial peptides, were produced likewise and verified to possess anti-microbial/anti-tumor bioactivities. No significant bacterial growth inhibition was observed during the fusion protein expression, indicating that the fusion form not only mediated the secretion but also decreased the toxicity of anti-microbial peptides (AMPs) to the host bacteria. To the best of our knowledge, this is the first report to achieve the secretory expression of these two AMPs in E. coli with considerable potential for manufacturing and industrialization purposes. Conclusions: The results demonstrate that the HlyA based novel production platform of E. coli allowed the efficient secretory production and purification of peptides, thus suggesting a promising strategy for the industrialized production of peptide pharmaceuticals or reagents. [ABSTRACT FROM AUTHOR]

Antibiotic resistance is rapidly increasing, and new anti-infective therapies are urgently needed. In this regard, antimicrobial peptides (AMPs) may represent potential candidates for the treatment of infections caused by multiresistant microorganisms. In this narrative review, we reported the experience of our research group over 20 years. We described the AMPs we evaluated against Gram-positive, Gram-negative, and fungi. In conclusion, our experience shows that AMPs can be a key option for treating multiresistant infections and overcoming resistance mechanisms. The combination of AMPs allows antibiotics and antifungals that are no longer effective to exploit the synergistic effect by restoring their efficacy. A current limitation includes poor data on human patients, the cost of some AMPs, and their safety, which is why studies on humans are needed as soon as possible.

Acanthamoeba is a free-living protozoan known to cause keratitis most commonly, especially among contact lens wearers. Treatment of Acanthamoeba keratitis is challenging as Acanthamoeba can encyst from the active form, a trophozoite, into a hibernating cyst that is refractory to antibiotics and difficult to kill; therefore, there is a need for more effective anti-amoebic strategies. In this study, we have evaluated the anti-amoebic activity of the antimicrobial peptide mimic RK-758 against Acanthamoeba castellanii. RK-758 peptidomimetic was subjected to biological assays to investigate its amoebicidal, amoebistatic, anti-encystation, and anti-excystation effects on A. castellanii. The anti-amoebic activity of the peptide mimic RK-758 was compared with chlorhexidine against the Acanthamoeba castellanii ATCC30868 and Acanthamoeba castellanii 044 (a clinical strain) with the concentrations of both ranging from 125 µM down to 7.81 µM. All experiments were performed in duplicate with three independent replicates. The data were represented as mean ± SE and analysed using a two-sample t-test and two-tailed distributions. A p < 0.05 was considered statistically significant. The peptidomimetic RK-758 had anti-Acanthamoeba activity against both trophozoites and cysts in a dose-dependent manner. The RK-758 had amoebicidal and growth inhibitory activities of ≥50% at a concentration between 125 µM and 15.6 µM against the trophozoites of both Acanthamoeba strains. Inhibitory effects on the cyst formation and trophozoite re-emergence from cysts were noted at similar concentrations. Chlorhexidine had 50% activity at 7.81 µM and above against the trophozoites and cysts of both strains. In the haemolysis assay, the RK-758 lysed horse RBCs at concentrations greater than 50 µM whereas lysis occurred at concentrations greater than 125 µM for the chlorhexidine. The peptidomimetic RK-758, therefore, has activity against both the trophozoite and cyst forms of Acanthamoeba and has the potential to be further developed as an anti-microbial agent against Acanthamoeba. RK-758 may also have use as an anti-amoebic disinfectant in contact lens solutions.

The skin secretions of many frogs have genetically-encoded, endogenous antimicrobial peptides (AMPs). Other species, especially aposematic poison frogs, secrete exogenously derived alkaloids that serve as potent defense molecules. The origins of these defense systems are not clear, but a novel bile-acid derived metabolite, tauromantellic acid, was recently discovered and shown to be endogenous in poison frogs (Mantella, Dendrobates, and Epipedobates). These observations raise questions about the evolutionary history of AMP genetic elements, the mechanism and function of tauromatellic acid production, and links between these systems. To understand the diversity and expression of AMPs among frogs, we assembled skin transcriptomes of 13 species across the anuran phylogeny. Our analyses revealed a diversity of AMPs and AMP expression levels across the phylogenetic history of frogs, but no observations of AMPs in Mantella. We examined genes expressed in the bile-acid metabolic pathway and found that CYP7A1 (Cytochrome P450), BAAT (bile acid-CoA: amino acid N-acyltransferase), and AMACR (alpha-methylacyl-CoA racemase) were highly expressed in the skin of M. betsileo and either lowly expressed or absent in other frog species. In particular, CYP7A1 catalyzes the first reaction in the cholesterol catabolic pathway and is the rate-limiting step in regulation of bile acid synthesis, suggesting unique activation of the bile acid pathway in Mantella skin. The activation of the bile acid pathway in the skin of Mantella and the lack of observed AMPs fuel new questions about the evolution of defense compounds and the ectopic expression of the bile-acid pathway.

The rise of antibiotic resistant bacteria due to overuse and misuse of antibiotics in medicine and dentistry is a growing concern. New approaches are needed to combat antibiotic resistant (AR) bacterial infections. There are a number of methods available and in development to address AR infections. Dentists conventionally use chemicals such as chlorohexidine and calcium hydroxide to kill oral bacteria, with many groups recently developing more biocompatible antimicrobial peptides (AMPs) for use in the oral cavity. AMPs are promising candidates in the treatment of (oral) infections. Also known as host defense peptides, AMPs have been isolated from animals across all kingdoms of life and play an integral role in the innate immunity of both prokaryotic and eukaryotic organisms by responding to pathogens. Despite progress over the last four decades, there are only a few AMPs approved for clinical use. This review summarizes an Introduction to Oral Microbiome and Oral Infections, Traditional Antibiotics and Alternatives & Antimicrobial Peptides. There is a focus on cationic AMP characteristics and mechanisms of actions, and an overview of animal-derived natural and synthetic AMPs, as well as observed microbial resistance.

Diabetic hyperglycemia delays wound healing, leading to serious consequences. Topical antibiotics can reduce the risk of a wound infection during healing; nevertheless, the microbial fight against antibiotics brings about public health challenges. Anti-microbial peptides (AMPs) belong to a novel class of drug that is used to prevent and treat systemic and topical infections. The aim of the current work was to achieve better wound healing in diabetic rats by conjugating the anti-microbial peptide “apamin” (APA) with the broad-spectrum antibiotic “ceftriaxone” (CTX) to form a nanocomplex. The CTX–APA nanoconjugate formulation was optimized using a Box–Behnken design. The optimized CTX–APA nanoconjugate formulation was evaluated for its size and zeta potential, and was then examined using transmission electron microscopy (TEM). The CTX–APA nanoconjugate was loaded onto a hydroxypropyl methylcellulose (2% w/v)-based hydrogel. It was observed that the application of the CTX–APA nanocomplex on the wounded skin of diabetic rats accelerated the regeneration of the epithelium, granulation tissue formation, epidermal proliferation, and keratinization. The nanocomplex was capable of significantly reducing the expression of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6), while increasing the expression of transforming growth factor beta-1 (TGF-β1) as well as the angiogenic markers: hypoxia-inducible factor 1-alpha (HIF-1α) and vascular endothelial growth factor (VEGF). Conclusively, the application of an ion-paired CTX–APA nanocomplex enhances wound healing in diabetic rats.

Objectives: Anti‐microbial peptides (AMPs) have been comprehensively investigated as a novel alternative to traditional antibiotics against microorganisms. Meanwhile, Tetrahedral DNA nanostructures (TDNs) have gained attention in the field of biomedicine for their premium biological effects and transportation efficiency as delivery vehicles. Hence, in this study, TDN/Histatin 5 (His‐5) was synthesized and the transport efficiency and anti‐fungal effect were measured to evaluate the promotion of His‐5 modified by TDNs. Materials and Methods: Tetrahedral DNA nanostructures/His‐5 complex was prepared via electrostatic attraction and characterized by transmission electron microscopy (TEM), polyacrylamide gel electrophoresis (PAGE), dynamic light scattering (DLS) and electrophoretic light scattering (ELS). The anti‐fungal effect of the TDN/His‐5 complex was evaluated by determining the growth curve and colony‐forming units of C. albicans. The morphological transformation of C. albicans was observed by light microscope and scanning electron microscope (SEM). Immunofluorescence was performed, and potassium efflux was detected to mechanistically demonstrate the efficacy of TDN/His‐5. Results: The results showed that Histatin 5 modified by TDNs had preferable stability in serum and was effectively transported into C. albicans, leading to the increased formation of intracellular reactive oxygen species, higher potassium efflux and enhanced anti‐fungal effect against C. albicans. Conclusions: Our study showed that TDN/His‐5 was synthesized successfully. And by the modification of TDNs, His‐5 showed increased transport efficiency and improved anti‐fungal effect. [ABSTRACT FROM AUTHOR]

Low body mass index (BMI) is a risk factor for progression from latent Mycobacterium tuberculosis infection to active tuberculosis (TB) disease. Anti-microbial peptides (AMPs) are multifunctional molecules that play a crucial role in the mammalian host innate defense mechanism. AMPs have been shown to have an important role in host immunity to TB infection. The association of antimicrobial peptides with low BMI–latent tuberculosis (LTBI) co-morbidity has not been explored. To study the association of AMPs with LTBI-BMI, we examined the systemic, baseline, and mycobacterial antigen stimulated levels of human neutrophil peptides 1–3, (HNP1-3), granulysin, human beta defensin–2 (HBD-2), and cathelicidin (LL-37) in individuals with LTBI and low BMI (LBMI) and compared them with individuals with LTBI and normal BMI (NBMI). LBMI was characterized by diminished systemic levels of HNP1-3, granulysin, HBD-2 and cathelicidin in comparison with NBMI. Similarly, LBMI was also characterized by diminished unstimulated levels of HNP1-3 and granulysin and diminished mycobacterial antigen stimulated levels of HNP1-3, granulysin, and HBD-2. In addition, certain AMPs exhibited a positive correlation with BMI. Our data, therefore, demonstrates that coexistent LBMI in LTBI is characterized by the diminished levels of HNP1-3, granulysin, HBD-2, and cathelicidin, thereby potentially increasing the risk of progression to active TB.

Filamentous fungi are a large and diverse taxonomically group of microorganisms found in all habitats worldwide. They grow as a network of cells called hyphae. Since filamentous fungi live in very diverse habitats, they produce different enzymes to degrade material for their living, for example hydrolytic enzymes to degrade various kinds of biomasses. Moreover, they produce defense proteins (antimicrobial peptides) and proteins for attaching surfaces (hydrophobins). Many of them are easy to cultivate in different known setups (submerged fermentation and solid-state fermentation) and their secretion of proteins and enzymes are often much larger than what is seen from yeast and bacteria. Therefore, filamentous fungi are in many industries the preferred production hosts of different proteins and enzymes. Edible fungi have traditionally been used as food, such as mushrooms or in fermented foods. New trends are to use edible fungi to produce myco-protein enriched foods. This review gives an overview of the different kinds of proteins, enzymes, and peptides produced by the most well-known fungi used as cell factories for different purposes and applications. Moreover, we describe some of the challenges that are important to consider when filamentous fungi are optimized as efficient cell factories.

Low body mass index (BMI) is a risk factor for progression from latent Mycobacterium tuberculosis infection to active tuberculosis (TB) disease. Anti-microbial peptides (AMPs) are multifunctional molecules that play a crucial role in the mammalian host innate defense mechanism. AMPs have been shown to have an important role in host immunity to TB infection. The association of antimicrobial peptides with low BMI–latent tuberculosis (LTBI) co-morbidity has not been explored. To study the association of AMPs with LTBI-BMI, we examined the systemic, baseline, and mycobacterial antigen stimulated levels of human neutrophil peptides 1–3, (HNP1-3), granulysin, human beta defensin–2 (HBD-2), and cathelicidin (LL-37) in individuals with LTBI and low BMI (LBMI) and compared them with individuals with LTBI and normal BMI (NBMI). LBMI was characterized by diminished systemic levels of HNP1-3, granulysin, HBD-2 and cathelicidin in comparison with NBMI. Similarly, LBMI was also characterized by diminished unstimulated levels of HNP1-3 and granulysin and diminished mycobacterial antigen stimulated levels of HNP1-3, granulysin, and HBD-2. In addition, certain AMPs exhibited a positive correlation with BMI. Our data, therefore, demonstrates that coexistent LBMI in LTBI is characterized by the diminished levels of HNP1-3, granulysin, HBD-2, and cathelicidin, thereby potentially increasing the risk of progression to active TB. [ABSTRACT FROM AUTHOR]

Cryptosporidium spp. are enteric protozoa parasites that infect a variety of vertebrate hosts. These parasites are capable of inducing life-threatening gastrointestinal disease in immunocompromised individuals. With the rising epidemiological evidence of the occurrence of Cryptosporidium infections in humans with digestive cancer, the tumorigenic potential of the parasite has been speculated. In this regard, Cryptosporidium parvum has been reported to induce digestive adenocarcinoma in a rodent model of chronic cryptosporidiosis. However, the processes by which the parasite could induce this carcinogenesis are still unknown. Therefore, the transcriptomes of C. parvum infected ileo-cecal regions of mice developing tumors were analyzed in the current study. For the first time, downregulation of the expression of α-defensin, an anti-microbial target of the parasite in response to C. parvum infection was observed in the transformed tissues. This phenomenon has been speculated to be the result of resistance of C. parvum to the host defense through the upregulated expression of interferon γ-stimulated genes. The inflammatory response generated as result of attenuated expression of anti-microbial peptides highlights the role of immune evasion in the C. parvum-induced tumorigenesis. The study has also succeeded in the characterization of the tumor microenvironment (TME) which is characterized by the presence of cancer associated fibroblasts, myeloid-derived suppressor cells, tumor-associated macrophages and extracellular matrix components. Identification of immune suppressor cells and accumulation of pro-inflammatory mediators speculates that chronic inflammation induced by persistent C. parvum infection assists in development of an immunosuppressive tumor microenvironment.

With rapidly ageing populations, the world is experiencing unsustainable healthcare from chronic diseases such as metabolic, cardiovascular, neurodegenerative, and cancer disorders. Healthy diet and lifestyle might contribute to prevent these diseases and potentially enhance health outcomes in patients during and after therapy. Fermented dairy foods (FDFs) found their origin concurrently with human civilization for increasing milk shelf-life and enhancing sensorial attributes. Although the probiotic concept has been developed more recently, FDFs, such as milks and yoghurt, have been unconsciously associated with health-promoting effects since ancient times. These health benefits rely not only on the occurrence of fermentation-associated live microbes (mainly lactic acid bacteria; LAB), but also on the pro-health molecules (PHMs) mostly derived from microbial conversion of food compounds. Therefore, there is a renaissance of interest toward traditional fermented food as a reservoir of novel microbes producing PHMs, and "hyperfoods" can be tailored to deliver these healthy molecules to humans. In FDFs, the main PHMs are bioactive peptides (BPs) released from milk proteins by microbial proteolysis. BPs display a pattern of biofunctions such as anti-hypertensive, antioxidant, immuno-modulatory, and anti-microbial activities. Here, we summarized the BPs most frequently encountered in dairy food and their biological activities; we reviewed the main studies exploring the potential of dairy microbiota to release BPs; and delineated the main effectors of the proteolytic LAB systems responsible for BPs release. [ABSTRACT FROM AUTHOR]

The skin secretions of many frogs have genetically-encoded, endogenous antimicrobial peptides (AMPs). Other species, especially aposematic poison frogs, secrete exogenously derived alkaloids that serve as potent defense molecules. The origins of these defense systems are not clear, but a novel bileacid derived metabolite, tauromantellic acid, was recently discovered and shown to be endogenous in poison frogs (Mantella, Dendrobates, and Epipedobates). These observations raise questions about the evolutionary history of AMP genetic elements, the mechanism and function of tauromatellic acid production, and links between these systems. To understand the diversity and expression of AMPs among frogs, we assembled skin transcriptomes of 13 species across the anuran phylogeny. Our analyses revealed a diversity of AMPs and AMP expression levels across the phylogenetic history of frogs, but no observations of AMPs in Mantella. We examined genes expressed in the bile-acid metabolic pathway and found that CYP7A1 (Cytochrome P450), BAAT (bile acid-CoA: amino acid N-acyltransferase), and AMACR (alphamethylacyl- CoA racemase) were highly expressed in the skin of M. betsileo and either lowly expressed or absent in other frog species. In particular, CYP7A1 catalyzes the first reaction in the cholesterol catabolic pathway and is the rate-limiting step in regulation of bile acid synthesis, suggesting unique activation of the bile acid pathway in Mantella skin. The activation of the bile acid pathway in the skin of Mantella and the lack of observed AMPs fuel new questions about the evolution of defense compounds and the ectopic expression of the bile-acid pathway. [ABSTRACT FROM AUTHOR]

Muscle contraction brings about movement and locomotion in animals. However, muscles have also been implicated in several atypical physiological processes including immune response. The role of muscles in immunity and the mechanism involved has not yet been deciphered. In this paper, using Drosophila indirect flight muscles (IFMs) as a model, we show that muscles are immune-responsive tissues. Flies with defective IFMs are incapable of mounting a potent humoral immune response. Upon immune challenge, the IFMs produce anti-microbial peptides (AMPs) through the activation of canonical signaling pathways, and these IFM-synthesized AMPs are essential for survival upon infection. The trunk muscles of zebrafish, a vertebrate model system, also possess the capacity to mount an immune response against bacterial infections, thus establishing that immune responsiveness of muscles is evolutionarily conserved. Our results suggest that physiologically fit muscles might boost the innate immune response of an individual.

Within a series of dipeptide derivatives (5–11), compound 4 was refluxed with d-glucose, d-xylose, acetylacetone, diethylmalonate, carbon disulfide, ethyl cyanoacetate, and ethyl acetoacetate which yielded 5–11, respectively. The candidates 5–11 were characterized and their biological activities were evaluated where they showed different anti-microbial inhibitory activities based on the type of pathogenic microorganisms. Moreover, to understand modes of binding, molecular docking was used of Nicotinoylglycine derivatives with the active site of the penicillin-binding protein 3 (PBP3) and sterol 14-alpha demethylase’s (CYP51), and the results, which were achieved via covalent and non-covalent docking, were harmonized with the biological activity results. Therefore, it was extrapolated that compounds 4, 7, 8, 9, and 10 had good potential to inhibit sterol 14-alpha demethylase and penicillin-binding protein 3; consequently, these compounds are possibly suitable for the development of a novel antibacterial and antifungal therapeutic drug. In addition, in silico properties of absorption, distribution, metabolism, and excretion (ADME) indicated drug likeness with low to very low oral absorption in most compounds, and undefined blood–brain barrier permeability in all compounds. Furthermore, toxicity (TOPKAT) prediction showed probability values for all carcinogenicity models were medium to pretty low for all compounds.

Toll receptors are involved in innate immunity of invertebrates. In this study, we identify and characterize two Toll genes (named HcToll4 and HcToll5) from triangle sail mussel Hyriopsis cumingii. HcToll4 has complete cDNA sequence of 3,162 bp and encodes a protein of 909 amino acids. HcToll5 cDNA is 4,501 bp in length and encodes a protein of 924 amino acids. Both deduced HcToll4 and HcToll5 protein contain signal peptide, extracellular leucine rich repeats (LRRs), and intracellular Toll/interleukin-1 (IL-1) receptor domains. Quantitative real-time PCR analysis revealed that HcToll4 and HcToll5 were largely distributed in the hepatopancreas and could be detected in the gills and mantle. HcToll4 and HcToll5 expression could respond to Staphylococcus aureus, Vibrio parahaemolyticus, White spot syndrome virus (WSSV) or Poly I:C challenge. RNA interference by siRNA results showed that HcToll4 and HcToll5 were involved in the regulation of theromacin (HcThe) and whey acidic protein (HcWAP) expression. Based on these results, HcToll4 and HcToll5 might play pivotal function in H. cumingii innate immune response.

Host defense peptides (HDPs) are currently major focal points of medical research as infectious microbes are gaining resistance to existing drugs. They are effective against multi-drug resistant pathogens due to their unique primary target, biological membranes, and their peculiar mode of action. Even though HDPs from 60 Asian frog species belonging to 15 genera have been characterized, research into these peptides is at a very early stage. The purpose of this review is to showcase the status of peptide research in Asia. Here we provide a summary of HDPs from Asian frogs.

Background: The DNA-binding peptide LL37 is a suspected autoantigen in psoriasis. It can be found in neutrophil extracellular traps (NETs) which have been suggested to play a role in the pathogenesis of the disease. Citrullination, the conversion of peptidyl-arginine into peptidyl-citrulline, can be implicated in the formation of NETs. We hypothesized that citrullination increases LL37 immunogenicity and that NETs are a source of LL37., Objectives: We aimed to characterize cytokine responses of B cells and T cells to native and citrullinated LL37 (citLL37) and determine the prevalence and composition of circulating NETs in patients with psoriasis and healthy blood donors (HDs)., Methods: Mononuclear cells (MNCs) and serum were isolated from 20 HDs and 20 patients with psoriasis. The MNCs were stimulated with native LL37 and citLL37 and the proportion of cytokine-positive B cells and T cells was determined by flow cytometry. Circulating antibodies against native LL37 and citLL37 as well as circulating NETs were measured by ELISA, as was the content of LL37, citLL37, and IgG in the NETs., Results: CitLL37, but not native LL37, induced IFN-γ-production by T cells and B cells from psoriasis patients, as well as IL-10-production by the patients' CD4 + T cells. Serum from 40% of patients and 55% of HDs contained circulating NETs, of which 63% and 27%, respectively, contained LL37. Only two patients had NETs containing citLL37 and IgG antibodies were found in NETs from three patients and one HD. Post-hoc analysis of the cytokines produced by B cells and T cells after stimulation with citLL37 revealed two clusters of patients consisting of 10 high-responders and 9 low-responders. The high-responders were those that had circulating NETs in combination with an earlier age of onset of the disease., Conclusion: Citrullinated but not native LL37 elicits IFN-γ-responses by T cells and B cells from psoriasis patients, particularly those with circulating NETs and early disease onset, suggesting a role of citLL37 as an autoantigen in this subgroup of patients., Competing Interests: Outside of the submitted work: NL has been an honorary speaker for Eli Lilly, Janssen Cilag, and Sandoz. LS has received research funding from Novartis, Bristol-Myers Squibb, AbbVie, Janssen Pharmaceuticals, Sanofi, Almirall, the Danish National Psoriasis Foundation, the LEO Foundation and honoraria as consultant and/or speaker for AbbVie, Eli Lilly, Novartis, Pfizer, LEO Pharma, Janssen, UCB, Almirall, Bristol-Myers Squibb, and Sanofi. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Martín Monreal, Kvist-Hansen, Massarenti, Steffensen, Loft, Hansen, Ødum, Skov and Nielsen.)

Background: Infections caused by pathogenic microorganisms have increased the need for hospital care and have thus represented a public health problem and a significant financial burden. Classical treatments consisting of traditional antibiotics face several challenges today. Anti-microbial peptides (AMPs) are a conserved characteristic of the innate immune response among different animal species to defend against pathogenic microorganisms., Objectives: In this study, a new peptide sequence (mCHTL131-140) was designed using the in silico approach., Methods: Cathelicidin-2 (UniprotID: Q2IAL7) was used as a potential antimicrobial protein, and a novel 10 - 12 amino acids sequence AMP was designed using bioinformatics tools and the AMP databases. Then, the anti-bacterial, anti-biofilm, and anti-fungal properties of the peptide, as well as its hemolytic activity and cytotoxicity towards human fibroblast (HDF) cells, were investigated in vitro., Results: Online bioinformatics tools indicated that the peptide sequence could have anti-bacterial, anti-viral, anti-fungal, and anti-biofilm properties with little hemolytic properties. The experimental tests confirmed that mCHTL131-140 exhibited the best anti-bacterial properties against Acinetobacter baumannii and had fair anti-fungal properties. Besides, it did not cause red blood cell lysis and showed no cytotoxicity towards HDF cells., Conclusions: In general, the designed peptide can be considered a promising AMP to control hospital-acquired infections by A. baumannii ., Competing Interests: The authors declare that there is no conflict of interest., (Copyright © 2023, Fathi et al.)

Toll receptors are involved in innate immunity of invertebrates. In this study, we identify and characterize two Toll genes (named HcToll4 and HcToll5) from triangle sail mussel Hyriopsis cumingii. HcToll4 has complete cDNA sequence of 3,162 bp and encodes a protein of 909 amino acids. HcToll5 cDNA is 4,501 bp in length and encodes a protein of 924 amino acids. Both deduced HcToll4 and HcToll5 protein contain signal peptide, extracellular leucine rich repeats (LRRs), and intracellular Toll/interleukin-1 (IL-1) receptor domains. Quantitative real-time PCR analysis revealed that HcToll4 and HcToll5 were largely distributed in the hepatopancreas and could be detected in the gills and mantle. HcToll4 and HcToll5 expression could respond to Staphylococcus aureus, Vibrio parahaemolyticus, White spot syndrome virus (WSSV) or Poly I:C challenge. RNA interference by siRNA results showed that HcToll4 and HcToll5 were involved in the regulation of theromacin (HcThe) and whey acidic protein (HcWAP) expression. Based on these results, HcToll4 and HcToll5 might play pivotal function in H. cumingii innate immune response. [ABSTRACT FROM AUTHOR]

Cathelicidin peptide LL-37 plays an important role in the early host response against invading pathogens via its broad-spectrum anti-microbial activity. In this study, we investigated LL-37 expression in the inflamed mucosa of inflammatory bowel disease (IBD) patients. Furthermore, the regulatory mechanism of LL-37 induction was investigated in human colonic subepithelial myofibroblasts (SEMFs). LL-37 mRNA expression and protein secretion were analysed using real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Intracellular signalling pathways were analysed using immunoblotting and specific small interference RNA (siRNA). The expression of LL-37 mRNA was increased significantly in the inflamed mucosa of ulcerative colitis and Crohn's disease. The Toll-like receptor (TLR)-3 ligand, polyinosinic-polycytidylic acid (poly(I:C), induced LL-37 mRNA expression and stimulated LL-37 secretion in colonic SEMFs. The transfection of siRNAs specific for intracellular signalling proteins [Toll/IL-1R domain-containing adaptor-inducing interferon (IFN) (TRIF), tumour necrosis factor receptor-associated factor (TRAF)6, transforming growth factor β-activated kinase (TAK)1] suppressed the poly(I:C)-induced LL-37 mRNA expression significantly. Poly(I:C)-induced phosphorylation of mitogen-activated protein kinases (MAPKs) and activated nuclear factor kappa B (NF-κB) and activating factor protein (AP)-1. siRNAs specific for NF-κB and c-Jun inhibited poly(I:C)-induced LL-37 mRNA expression. LL-37 suppressed lipopolysaccharide (LPS)-induced interleukin (IL)-6 and IL-8 expression significantly in colonic SEMFs. The expression of LL-37 was up-regulated in the inflamed mucosa of IBD patients. LL-37 was induced by TLR-3 stimulation and exhibited an anti-microbial effect via interaction with lipopolysaccharide (LPS). [ABSTRACT FROM AUTHOR]

With rapidly ageing populations, the world is experiencing unsustainable healthcare from chronic diseases such as metabolic, cardiovascular, neurodegenerative, and cancer disorders. Healthy diet and lifestyle might contribute to prevent these diseases and potentially enhance health outcomes in patients during and after therapy. Fermented dairy foods (FDFs) found their origin concurrently with human civilization for increasing milk shelf-life and enhancing sensorial attributes. Although the probiotic concept has been developed more recently, FDFs, such as milks and yoghurt, have been unconsciously associated with health-promoting effects since ancient times. These health benefits rely not only on the occurrence of fermentation-associated live microbes (mainly lactic acid bacteria; LAB), but also on the pro-health molecules (PHMs) mostly derived from microbial conversion of food compounds. Therefore, there is a renaissance of interest toward traditional fermented food as a reservoir of novel microbes producing PHMs, and “hyperfoods” can be tailored to deliver these healthy molecules to humans. In FDFs, the main PHMs are bioactive peptides (BPs) released from milk proteins by microbial proteolysis. BPs display a pattern of biofunctions such as anti-hypertensive, antioxidant, immuno-modulatory, and anti-microbial activities. Here, we summarized the BPs most frequently encountered in dairy food and their biological activities; we reviewed the main studies exploring the potential of dairy microbiota to release BPs; and delineated the main effectors of the proteolytic LAB systems responsible for BPs release.

Using a combination of coarse-grained and atomistic molecular dynamics simulations we have investigated the membrane binding and folding properties of the membrane lytic peptide of Flock House virus (FHV). FHV is an animal virus and an excellent model system for studying cell entry mechanisms in non-enveloped viruses. FHV undergoes a maturation event where the 44 C-terminal amino acids are cleaved from the major capsid protein, forming the membrane lytic (γ) peptides. Under acidic conditions, γ is released from the capsid interior allowing the peptides to bind and disrupt membranes. The first 21 N-terminal residues of γ, termed γ 1 , have been resolved in the FHV capsid structure and γ 1 has been the subject of in vitro studies. γ 1 is structurally dynamic as it adopts helical secondary structure inside the capsid and on membranes, but it is disordered in solution. In vitro studies have shown the binding free energies to POPC or POPG membranes are nearly equivalent, but binding to POPC is enthalpically driven, while POPG binding is entropically driven. Through coarse-grained and multiple microsecond all-atom simulations the membrane binding and folding properties of γ 1 are investigated against homogeneous and heterogeneous bilayers to elucidate the dependence of the microenvironment on the structural properties of γ 1 . Our studies provide a rationale for the thermodynamic data and suggest binding of γ 1 to POPG bilayers occurs in a disordered state, but γ 1 must adopt a helical conformation when binding POPC bilayers. [ABSTRACT FROM AUTHOR]

Alpha helices (AH) are peptide fragments characterized by regular patterns of hydrogen bonding between the carbonyl oxygen and amino nitrogen of residues regularly spaced in sequence, resulting in spiral conformations. Their preponderance in protein structures underlines their importance. Interestingly, AHs are present in most anti-microbial peptides, although they might remain in random-coil conformations depending on the solvent dielectric. For example, the cecropin component of the chimeric anti-microbial protein designed previously by our group comprises of two AHs linked by a short stretch of random coil. These anti-microbial peptides are often amphipathic (quantified by a hydrophobic moment), aligning hydrophobic residues on one surface and charged residues on the others. In the current work, we reproduce previously described computational methods to compute the hydrophobic moment of AHs - and provide open access to the source code (PAGAL). We simultaneously generated input files for TikZ (a package for creating high resolution graphics programmatically) to obtain the Edmundson wheel and showing the direction and magnitude of the hydrophobic moment, and Pymol scripts to generate color coded protein surfaces. Additionally, we have observed an empirical structural property of AHs: the distance between the Cα atoms of the ith and (i+4)th residue is equal to the distance between the carbonyl oxygens of the ith and (i+4)th residue. We validated this using 100 non-homologous high resolution structures from the PISCES database. The source code and manual is available at http://github.com/sanchak/pagal and on http://dx.doi.org/10.5281/zenodo.11136.

This review summarises the current knowledge of Gomesin (Gm), an 18-residue long, cationic anti-microbial peptide originally isolated from the haemocytes of the Brazilian tarantula Acanthoscurria gomesiana. The peptide shows potent cytotoxic activity against clinically relevant microbes including Gram-positive and Gram-negative bacteria, fungi, and parasites. In addition, Gm shows in-vitro and in-vivo anti-cancer activities against several human and murine cancers. The peptide exerts its cytotoxic activity by permeabilising cell membranes, but the underlying molecular mechanism of action is still unclear. Due to its potential as a therapeutic agent, the structure and membrane-binding properties, as well as the leakage and cytotoxic activities of Gm have been studied using a range of techniques. This review provides a summary of these studies, with a particular focus on biophysical characterisation studies of peptide variants that have attempted to establish a structure-activity relationship. Future studies are still needed to rationalise the binding affinity and cell-type-specific selectivity of Gm and its variants, while more pre-clinical studies are required to develop Gm into a therapeutically useful peptide.

Background: In preclinical models of Type 1 Diabetes (T1D) the integrity of the gut barrier (GB) is instrumental to avoid dysregulated crosstalk between the commensal microbiota and immune cells and to prevent autoimmunity. The GB is composed of the intestinal epithelial barrier (IEB) and of the mucus layer containing mucins and antimicrobial peptides (AMPs) that are crucial to maintain immune tolerance. In preclinical models of T1D the alterations of the GB primarily affect the mucus layer. In human T1D increased gut permeability and IEB damage have been demonstrated but the integrity of the mucus layer was never assessed., Methods: We evaluated GB integrity by measuring serological markers of IEB damage (serological levels of zonulin) and bacterial translocation such as lipopolysaccharide binding protein (LBP) and myeloid differentiation protein 2 (MD2), and mRNA expression of tight junction proteins, mucins and AMPs in intestinal tissue of T1D patients and healthy controls (HC). Simultaneously, we performed immunological profiling on intestinal tissue and 16S rRNA analysis on the mucus-associated gut microbiota (MAGM)., Findings: Our data show a GB damage with mucus layer alterations and reduced mRNA expression of several mucins (MUC2, MUC12, MUC13, MUC15, MUC20, MUC21) and AMPs (HD4 and HD5) in T1D patients. Mucus layer alterations correlated with reduced relative abundance of short chain fatty acids (SCFA)-producing bacteria such as Bifidobacterium dentium, Clostridium butyricum and Roseburia intestinalis that regulate mucin expression and intestinal immune homeostasis. In T1D patients we also found intestinal immune dysregulation with higher percentages of effector T cells such as T helper (Th) 1, Th17 and TNF-α + T cells., Interpretation: Our data show that mucus layer alterations are present in T1D subjects and associated with dysbiosis and immune dysregulation., Funding: Research Grants from the Juvenile Diabetes Foundation (Grant 1-INO-2018-640-A-N to MF and 2-SRA-2019-680-S-B to JD) and from the Italian Ministry of Health (Grant RF19-12370721 to MF)., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Bacteria produce and excrete a versatile and dynamic suit of compounds to defend against microbial competitors and mediate local population dynamics. These include a wide range of broad-spectrum non-ribosomally synthesized antibiotics, lytic enzymes, metabolic by-products, proteinaceous exotoxins and ribosomally produced antimicrobial peptides (bacteriocins). Most bacteria produce at least one bacteriocin. Bacteriocins are of interest in the food industry as natural preservatives and in the probiotics industry, leading to extensive studies on lactic acid bacteria (colicin produced by Escherichia coli is a model bacteriocin). Recent studies have projected use of bacteriocins in veterinary medicine and in agriculture, as a biostimulants of plant growth and development and as biocontrol agents. For example, bacteriocins such as Cerein 8A, Bac-GM17, putidacin, Bac 14B, amylocyclicin have been studied for their mechanisms of anti-microbial activity. Bac IH7 promotes tomato and musk melon plant growth. Thuricin 17 (Th17) is the only bacteriocin studied extensively for plant growth promotion and at the molecular level. Th17 functions as a bacterial signal compound, promoting plant growth in legumes and non-legumes. In Arabidopsis thaliana and Glycine max Th17 increased phytohormones IAA and SA at 24 h post treatment. At the proteome level Th17 treatment of 3-week-old A. thaliana rosettes led to > 2-fold changes in activation of the carbon and energy metabolism pathway proteins, 24 h post treatment. At 250 mM NaCl stress, the control plants under osmotic-shock shut down most of carbon-metabolism and activated energy-metabolism and antioxidant pathways. Th17 treated plants, at 250 mM NaCl, retained meaningful levels of the light harvesting complex, photosystem I and II proteins and energy and antioxidant pathways were activated, so that rosettes could better withstand the salt stress. In Glycine max, Th17 helped seeds germinate in the presence of NaCl stress, and was most effective at 100 mM NaCl. The 48 h post germination proteome suggested efficient and speedier partitioning of storage proteins, activation of carbon, nitrogen and energy metabolisms in Th17 treated seeds both under optimal and 100 mM NaCl. This review focuses on the bacteriocins produced by plant-rhizosphere colonizers and plant-pathogenic bacteria, that might have uses in agriculture, veterinary and human medicine.

We have observed previously that micronutrient supplementation ameliorated suppression of α-defensin expression during diarrhoea. However, how interactions between anti-microbial peptide (AMP) expression and diarrhoeal disease are altered by micronutrient supplementation remain unclear. Using oral vaccination as a model of intestinal infection, we measured changes in AMP expression during multiple micronutrient supplementation. In the first part, volunteers underwent duodenal jejunal biopsy before and at 1, 2, 4 or 7 days after administration of one of three live, attenuated oral vaccines against rotavirus, typhoid and enterotoxigenic Escherichia coli. In the second part, participants were randomized to receive a multiple micronutrient supplement or placebo for 6 weeks before undergoing intestinal biopsy, vaccination against typhoid and rebiopsy after 14 days. Expression of human alpha-defensin (HD)5, HD6, hBD1, hBD2 and LL-37 was measured by quantitative reverse transcription-polymerase chain reaction. Taken together, the bacterial vaccines, but not rotavirus vaccine, reduced HD5 expression ( P = 0·02, signed-rank test) and reduced LL-37 expression in seven of the eight individuals whose biopsies had expression prevaccination ( P = 0·03). hBD2 was not detected. In the controlled trial, HD5 and HD6 expression after vaccination was lower [median ratio 0·5, interquartile range (IQR) = 0·07-2·2 and 0·58, IQR = 0·13-2·3, respectively] than before vaccination. There was no significant effect detected of micronutrient supplementation on expression of HD5, HD6, hBD1 or LL-37. We conclude that live attenuated bacterial vaccines, but not rotavirus vaccine, can reduce intestinal α-defensins, and typhoid vaccine reduced LL-37 expression. We found no evidence that micronutrient supplementation in the short term had any impact on anti-microbial peptide expression. [ABSTRACT FROM AUTHOR]