Gut Susceptibility to Viral Invasion: Contributing Roles of Diet, Microbiota and Enteric Nervous System to Mucosal Barrier Preservation.

The gastrointestinal lumen is a rich source of eukaryotic and prokaryotic viruses which, together with bacteria, fungi and other microorganisms comprise the gut microbiota. Pathogenic viruses inhabiting this niche have the potential to induce local as well as systemic complications; among them, the viral ability to disrupt the mucosal barrier is one mechanism associated with the promotion of diarrhea and tissue invasion. This review gathers recent evidence showing the contributing effects of diet, gut microbiota and the enteric nervous system to either support or impair the mucosal barrier in the context of viral attack.

The Immune Characteristics of the Epididymis and the Immune Pathway of the Epididymitis Caused by Different Pathogens.

The epididymis is an important male accessory sex organ where sperm motility and fertilization ability develop. When spermatozoa carrying foreign antigens enter the epididymis, the epididymis shows "immune privilege" to tolerate them. It is well-known that a tolerogenic environment exists in the caput epididymis, while pro-inflammatory circumstances prefer the cauda epididymis. This meticulously regulated immune environment not only protects spermatozoa from autoimmunity but also defends spermatozoa against pathogenic damage. Epididymitis is one of the common causes of male infertility. Up to 40% of patients suffer from permanent oligospermia or azoospermia. This is related to the immune characteristics of the epididymis itself. Moreover, epididymitis induced by different pathogenic microbial infections has different characteristics. This article elaborates on the distribution and immune response characteristics of epididymis immune cells, the role of epididymis epithelial cells (EECs), and the epididymis defense against different pathogenic infections (such as uropathogenic Escherichia coli, Chlamydia trachomatis, and viruses to provide therapeutic approaches for epididymitis and its subsequent fertility problems.

Aminoglycosides can be a better choice over macrolides in COVID-19 regimen: Plausible mechanism for repurposing strategy.

In the current COVID-19 pandemic, prioritizing the immunity enhancers is equally important to anti-virals. Defensins are the forgotten molecules that enhance the innate immunity against various microbes. Although macrolides like azithromycin and clarithromycin etc., have been reported to act against respiratory infections but they lack the ability of immunity enhancement through defensins. The aminoglycosides were proved to have defensin mediated antiviral activity, that could enhance the immunity. So, Consideration of aminoglycosides can be a double edge sword viz., against respiratory infection as well as Immunity enhancer (along with anti-virals) for COVID-19 regimen.

Functional Insights From the Evolutionary Diversification of Big Defensins.

Big defensins are antimicrobial polypeptides believed to be the ancestors of ß-defensins, the most evolutionary conserved family of host defense peptides (HDPs) in vertebrates. Nevertheless, big defensins underwent several independent gene loss events during animal evolution, being only retained in a limited number of phylogenetically distant invertebrates. Here, we explore the evolutionary history of this fascinating HDP family and investigate its patchy distribution in extant metazoans. We highlight the presence of big defensins in various classes of lophotrochozoans, as well as in a few arthropods and basal chordates (amphioxus), mostly adapted to life in marine environments. Bivalve mollusks often display an expanded repertoire of big defensin sequences, which appear to be the product of independent lineage-specific gene tandem duplications, followed by a rapid molecular diversification of newly acquired gene copies. This ongoing evolutionary process could underpin the simultaneous presence of canonical big defensins and non-canonical (ß-defensin-like) sequences in some species. The big defensin genes of mussels and oysters, two species target of in-depth studies, are subjected to gene presence/absence variation (PAV), i.e., they can be present or absent in the genomes of different individuals. Moreover, big defensins follow different patterns of gene expression within a given species and respond differently to microbial challenges, suggesting functional divergence. Consistently, current structural data show that big defensin sequence diversity affects the 3D structure and biophysical properties of these polypeptides. We discuss here the role of the N-terminal hydrophobic domain, lost during evolution toward ß-defensins, in the big defensin stability to high salt concentrations and its mechanism of action. Finally, we discuss the potential of big defensins as markers for animal health and for the nature-based design of novel therapeutics active at high salt concentrations.

Interactions of the Skin Pathogen Haemophilus ducreyi With the Human Host.

The obligate human pathogen Haemophilus ducreyi causes both cutaneous ulcers in children and sexually transmitted genital ulcers (chancroid) in adults. Pathogenesis is dependent on avoiding phagocytosis and exploiting the suppurative granuloma-like niche, which contains a myriad of innate immune cells and memory T cells. Despite this immune infiltrate, long-lived immune protection does not develop against repeated H. ducreyi infections-even with the same strain. Most of what we know about infectious skin diseases comes from naturally occurring infections and/or animal models; however, for H. ducreyi, this information comes from an experimental model of infection in human volunteers that was developed nearly three decades ago. The model mirrors the progression of natural disease and serves as a valuable tool to determine the composition of the immune cell infiltrate early in disease and to identify host and bacterial factors that are required for the establishment of infection and disease progression. Most recently, holistic investigation of the experimentally infected skin microenvironment using multiple "omics" techniques has revealed that non-canonical bacterial virulence factors, such as genes involved in central metabolism, may be relevant to disease progression. Thus, the immune system not only defends the host against H. ducreyi, but also dictates the nutrient availability for the invading bacteria, which must adapt their gene expression to exploit the inflammatory metabolic niche. These findings have broadened our view of the host-pathogen interaction network from considering only classical, effector-based virulence paradigms to include adaptations to the metabolic environment. How both host and bacterial factors interact to determine infection outcome is a current focus in the field. Here, we review what we have learned from experimental H. ducreyi infection about host-pathogen interactions, make comparisons to what is known for other skin pathogens, and discuss how novel technologies will deepen our understanding of this infection.

The Arabidopsis defensin gene AtPDF2.5 mediates cadmium tolerance and accumulation.

Although excess cadmium (Cd) accumulation is harmful to plants, the molecular mechanisms underlying Cd detoxification and accumulation in Arabidopsis thaliana remain largely undetermined. In this study, we demonstrated that the A. thaliana PLANT DEFENSIN 2 gene AtPDF2.5 is involved in Cd tolerance and accumulation. In vitro Cd-binding assays revealed that AtPDF2.5 has Cd-chelating activity. Site-directed mutagenesis of AtPDF2.5 identified eight cysteine residues that were essential for mediating Cd tolerance and chelation. Histochemical analysis demonstrated that AtPDF2.5 was mainly expressed in root xylem vascular bundles, and that AtPDF2.5 was significantly induced by Cd. Subcellular localization analysis revealed that AtPDF2.5 was localized to the cell wall. The overexpression of AtPDF2.5 significantly enhanced Cd tolerance and accumulation in A. thaliana and its heterologous overexpression in rice increased Cd accumulation; however, the functional disruption of AtPDF2.5 decreased Cd tolerance and accumulation. Physiological analysis suggested that AtPDF2.5 promoted Cd efflux from the protoplast and its subsequent accumulation in the cell wall. These data suggest that AtPDF2.5 promotes cytoplasmic Cd efflux via chelation, thereby enhancing Cd detoxification and apoplastic accumulation.

Antimicrobial proteins: intestinal guards to protect against liver disease.

Alterations of gut microbes play a role in the pathogenesis and progression of many disorders including liver and gastrointestinal diseases. Both qualitative and quantitative changes in gut microbiota have been associated with liver disease. Intestinal dysbiosis can disrupt the integrity of the intestinal barrier leading to pathological bacterial translocation and the initiation of an inflammatory response in the liver. In order to sustain symbiosis and protect from pathological bacterial translocation, antimicrobial proteins (AMPs) such as a-defensins and C-type lectins are expressed in the gastrointestinal tract. In this review, we provide an overview of the role of AMPs in different chronic liver disease such as alcoholic steatohepatitis, non-alcoholic fatty liver disease, and cirrhosis. In addition, potential approaches to modulate the function of AMPs and prevent bacterial translocation are discussed.

X-ray structure of a carpet-like antimicrobial defensin-phospholipid membrane disruption complex.

Defensins are cationic antimicrobial peptides expressed throughout the plant and animal kingdoms as a first line of defense against pathogens. Membrane targeting and disruption is a crucial function of many defensins, however the precise mechanism remains unclear. Certain plant defensins form dimers that specifically bind the membrane phospholipids phosphatidic acid (PA) and phosphatidylinositol 4,5-bisphosphate, thereby triggering the assembly of defensin-lipid oligomers that permeabilize cell membranes. To understand this permeabilization mechanism, here we determine the crystal structure of the plant defensin NaD1 bound to PA. The structure reveals a 20-mer that adopts a concave sheet- or carpet-like topology where NaD1 dimers form one face and PA acyl chains form the other face of the sheet. Furthermore, we show that Arg39 is critical for PA binding, oligomerization and fungal cell killing. These findings identify a putative defensin-phospholipid membrane attack configuration that supports a longstanding proposed carpet mode of membrane disruption.

Cell adhesion properties of human defensins.

Effector peptides of innate immunity play an important role in host defense. They act directly by inactivating microbes but also link innate to adaptive immunity. A variety of innate immune functions has been described for these peptides, including chemoattraction and cytokine release. In this study, we describe the effect on cell morphology and cell adhesion of human defensins. We find that Human Defensin 5, the major product of specialized gut epithelial cells, causes changes in cell morphology. HD-5 induces cell adhesion, binds to fibronectin and facilitates binding of T cells to intestinal epithelial cells. These effects were found also for a second prominent defensing, termed Human Neutrophil peptide-1, but not for other human defensins.

The mycorrhiza-dependent defensin MtDefMd1 of Medicago truncatula acts during the late restructuring stages of arbuscule-containing cells.

Different symbiotic and pathogenic plant-microbe interactions involve the production of cysteine-rich antimicrobial defensins. In Medicago truncatula, the expression of four MtDefMd genes, encoding arbuscular mycorrhiza-dependent defensins containing an N-terminal signal peptide and exhibiting some differences to non-symbiotic defensins, raised over the time of fungal colonization. Whereas the MtDefMd1 and MtDefMd2 promoters were inactive in cells containing young arbuscules, cells with fully developed arbuscules displayed different levels of promoter activities, indicating an up-regulation towards later stages of arbuscule formation. MtDefMd1 and MtDefMd2 expression was absent or strongly down-regulated in mycorrhized ram1-1 and pt4-2 mutants, known for defects in arbuscule branching or premature arbuscule degeneration, respectively. A ~97% knock-down of MtDefMd1/MtDefMd2 expression did not significantly affect arbuscule size. Although overexpression of MtDefMd1 in arbuscule-containing cells led to an up-regulation of MtRam1, encoding a key transcriptional regulator of arbuscule formation, no morphological changes were evident. Co-localization of an MtDefMd1-mGFP6 fusion with additional, subcellular markers revealed that this defensin is associated with arbuscules in later stages of their life-cycle. MtDefMd1-mGFP6 was detected in cells with older arbuscules about to collapse, and ultimately in vacuolar compartments. Comparisons with mycorrhized roots expressing a tonoplast marker indicated that MtDefMd1 acts during late restructuring processes of arbuscule-containing cells, upon their transition into a post-symbiotic state.

An anionic defensin from Plutella xylostella with potential activity against Bacillus thuringiensis.

Insect defensins, are cationic peptides that play an important role in immunity against microbial infection. In the present study, an anionic defensin from Plutella xylostella, (designated as PxDef) was first cloned and characterized. Amino acid sequence analysis showed that the mature peptide owned characteristic six-cysteine motifs with predicted isoelectric point of 5.57, indicating an anionic defensin. Quantitative real-time polymerase chain reaction analysis showed that PxDef was significantly induced in epidermis, fat body, midgut and hemocytes after injection of heat-inactivated Bacillus thuringiensis, while such an induction was delayed by the injection of live B. thuringiensis in the 4th instar larvae of P. xylostella. Knocking down the expression of nuclear transcription factor Dorsal in P. xylostella by RNA interference significantly decreased the mRNA level of PxDef, and increased the sensitivity of P. xylostella larvae to the infection by live B. thuringiensis. The purified recombinant mature peptide (PxDef) showed higher activity against Gram-positive bacteria, with the minimum inhibition concentrations of 1.6 and 2.6 µM against B. thuringiensis and Bacillus subtilis, respectively. To our knowledge, this is the first report about an anionic PxDef, which may play an important role in the immune system of P. xylostella against B. thuringiensis.

Antimicrobial potentials and structural disorder of human and animal defensins.

Defensins are moonlighting peptides which are broadly distributed throughout all the living kingdoms. They play a multitude of important roles in human health and disease, possessing several immunoregulatory functions and manifesting broad antimicrobial activities against viruses, bacteria, and fungi. Based on their patterns of intramolecular disulfide bridges, these small cysteine-rich cationic proteins are divided into three major types, α-, ß-, and θ-defensins, with the α- and ß-defensins being further subdivided into a number of subtypes. The various roles played by the defensins in the innate (especially mucosal) and adoptive immunities place these polypeptides at the frontiers of the defense against the microbial invasions. Current work analyzes the antimicrobial activities of human and animal defensins in light of their intrinsic disorder propensities.

Tribolium castaneum defensins are primarily active against Gram-positive bacteria.

The red flour beetle Tribolium castaneum is a destructive insect pest of stored food and feed products, and a model organism for development, evolutionary biology and immunity. The insect innate immune system includes antimicrobial peptides (AMPs) with a wide spectrum of targets including viruses, bacteria, fungi and parasites. Defensins are an evolutionarily-conserved class of AMPs and a potential new source of antimicrobial agents. In this context, we report the antimicrobial activity, phylogenetic and structural properties of three T. castaneum defensins (Def1, Def2 and Def3) and their relevance in the immunity of T. castaneum against bacterial pathogens. All three recombinant defensins showed bactericidal activity against Micrococcus luteus and Bacillus thuringiensis serovar tolworthi, but only Def1 and Def2 showed a bacteriostatic effect against Staphylococcus epidermidis. None of the defensins showed activity against the Gram-negative bacteria Escherichia coli and Pseudomonas entomophila or against the yeast Saccharomyces cerevisiae. All three defensins were transcriptionally upregulated following a bacterial challenge, suggesting a key role in the immunity of T. castaneum against bacterial pathogens. Phylogenetic analysis showed that defensins from T. castaneum, mealworms, Udo longhorn beetle and houseflies cluster within a well-defined clade of insect defensins. We conclude that T. castaneum defensins are primarily active against Gram-positive bacteria and that other AMPs may play a more prominent role against Gram-negative species.

Functional synergism of Human Defensin 5 and Human Defensin 6.

The gut epithelium is critically involved in maintaining intestinal immune homeostasis. Acting as a physical barrier, it separates the intestinal microflora from cells of the immune system. In addition to its barrier function, the intestinal epithelium expresses defensins, natural, endogenous antimicrobial peptides. In humans, specialized epithelial cells, termed Paneth cells, located primarily in the small intestine express two defensins, Human Defensin-5 (HD-5) and Human Defensin-6 (HD-6). Previously, we have shown that HD-5 potently kills bacteria and induces secretion of interleukin-8 by intestinal epithelial cells. We show that HD-6 specifically and synergistically enhances the HD-5-induced IL-8 secretion, but does not alter its anti-bacterial activity. Further, we find that HD-5 decreases the trans-epithelial electrical resistance of intestinal epithelial cells and that HD-6 negates this effect of HD-5.

Diverse functions of defensins and other antimicrobial peptides in periodontal tissues.

Defensins are antimicrobial peptides that exhibit direct microbicidal activity as well as mediator-like functions by, for example, activating immature dendritic cells. This review focuses on defensins and other antimicrobial peptides that are present in periodontal tissues. Their antimicrobial capacity against periodontal microorganisms, their regulation and their expression profiles during periodontal diseases is discussed. As antimicrobial peptides may possess great potential for new diagnostic, preventive and therapeutic strategies, a better understanding of how antimicrobial peptides are regulated as part of the innate host immune response is crucial.

Amplifying renal immunity: the role of antimicrobial peptides in pyelonephritis.

Urinary tract infections (UTIs), including pyelonephritis, are among the most common and serious infections encountered in nephrology practice. UTI risk is increased in selected patient populations with renal and urinary tract disorders. As the prevalence of antibiotic-resistant uropathogens increases, novel and alternative treatment options will be needed to reduce UTI-associated morbidity. Discoveries over the past decade demonstrate a fundamental role for the innate immune system in protecting the urothelium from bacterial challenge. Antimicrobial peptides, an integral component of this urothelial innate immune system, demonstrate potent bactericidal activity toward uropathogens and might represent a novel class of UTI therapeutics. The urothelium of the bladder and the renal epithelium secrete antimicrobial peptides into the urinary stream. In the kidney, intercalated cells--a cell-type involved in acid-base homeostasis--have been shown to be an important source of antimicrobial peptides. Intercalated cells have therefore become the focus of new investigations to explore their function during pyelonephritis and their role in maintaining urinary tract sterility. This Review provides an overview of UTI pathogenesis in the upper and lower urinary tract. We describe the role of intercalated cells and the innate immune response in preventing UTI, specifically highlighting the role of antimicrobial peptides in maintaining urinary tract sterility.

The first anionic defensin from amphibians.

A variety of antimicrobial peptides against infection have been identified from the skin of amphibians. However, knowledge on amphibian defensins is very limited. A novel anionic defensin designated PopuDef was purified from the skin of tree frog Polypedates puerensis, and the cDNA encoding PopuDef precursor was cloned from the skin cDNA library. The amino acid sequence of PopuDef (net charge: -2, pI: 4.75) shared the highest identity of 57 % (25/44) with the salamander defensin CFBD-1 (net charge: 0, pI: 6.14) from urodela amphibians. PopuDef showed moderate antimicrobial activities against P. aeruginosa and S. aureus (MICs are 19.41 and 17.25 µM, respectively), and relatively weak activities against E. coli and B. subtilis (MICs are 38.82 and 43.14 µM, respectively). Tissue distribution analysis indicated that relatively high expression level of PopuDef mRNA was observed in immune-related tissues including skin, gut, lung and spleen. Furthermore, the expression level of PopuDef was significantly upregulated in these tissues after tree frogs were infected with different bacteria strains mentioned above. Interestingly, the induction of PopuDef challenged with E. coli or B. subtilis, which was less sensitive to PopuDef, was much higher than that did with P. aeruginosa or S. aureus. These findings highlight the key role of PopuDef in innate immunity against infection. To our knowledge, PopuDef is the first anionic defensin characterized from amphibians.

Antifungal plant defensins: mechanisms of action and production.

Plant defensins are small, cysteine-rich peptides that possess biological activity towards a broad range of organisms. Their activity is primarily directed against fungi, but bactericidal and insecticidal actions have also been reported. The mode of action of various antifungal plant defensins has been studied extensively during the last decades and several of their fungal targets have been identified to date. This review summarizes the mechanism of action of well-characterized antifungal plant defensins, including RsAFP2, MsDef1, MtDef4, NaD1 and Psd1, and points out the variety by which antifungal plant defensins affect microbial cell viability. Furthermore, this review summarizes production routes for plant defensins, either via heterologous expression or chemical synthesis. As plant defensins are generally considered non-toxic for plant and mammalian cells, they are regarded as attractive candidates for further development into novel antimicrobial agents.