Antimicrobial Peptides/proteins Research Articles

Epithelial Antimicrobial Peptide/Protein and Cytokine Expression Profiles Obtained from Nasopharyngeal Swabs of SARS-CoV-2-Infected and Non-Infected Subjects.

Immune responses of the epithelia of the upper respiratory tract are likely crucial in early inhibition of the viral replication and finally clearance of SARS-CoV-2. We aimed to compare the expression profiles of antimicrobial peptides/proteins (AMPs) and related cytokines observed in the nasopharynx of SARS-CoV-2-infected patients and non-infected controls and to assess the associations between these parameters and COVID-19 patients' outcomes. We included 45 subjects who had tested positive for SARS-CoV-2 and 22 control subjects who had tested negative for SARS-CoV-2. Biomaterial for SARS-CoV-2 detection, as well as gene and protein expression studies, was obtained from all subjects using nasopharyngeal swabs which were performed a maximum of 7 days before inclusion in the study. Univariable and multivariable statistics were performed. When compared to the controls, the mRNA expression levels of human β-defensin 1 (hBD-1), LL-37, and trappin-2 were significantly higher in specimens of nasopharyngeal swabs from COVID-19 patients. Protein expression of hBD-1 was also increased in the COVID-19 group. mRNA expression levels of interferon-ɣ (IFN-ɣ), tumor necrosis factor- ɑ (TNF-ɑ), and interleukin-6 (IL-6) measured in SARS-CoV-2-infected patients were significantly higher than those observed in the controls, which could also be confirmed in the protein levels of IFN-ɣ and IL-6. A significant correlation between mRNA and protein levels could be observed only for IL-6. Univariable analysis revealed that low IFN-ɣ mRNA levels were associated with severe/fatal outcomes. The occurrence of COVID-19 pneumonia was significantly associated with lower expression levels of IL-6 mRNA, IFN-ɣ mRNA, and TNF-ɑ mRNA. Concerning the severe/fatal outcomes, the multivariable logistic regression model revealed that none of the aforementioned parameters remained significant in the model. However, the logistic regression model revealed that higher TNF-ɑ mRNA expression was a significant independent predictor of absence of pneumonia [odds ratio: 0.35 (95% CI 0.14 to 0.88, p = 0.024)]. In conclusion, nasopharyngeal expression of AMPs (hBD-1, LL-37, and trappin-2) and cytokines (IL-6, IFN-ɣ, and TNF-ɑ) is upregulated in response to early SARS-CoV-2 infection, indicating that these AMPs and cytokines play a role in the local host defense against the virus. Upregulated nasopharyngeal TNF-ɑ mRNA expression during the early phase of SARS-CoV-2 infection was a significant independent predictor of the absence of COVID-19 pneumonia. Hence, high TNF-ɑ mRNA expression in the nasopharynx appears to be a protective factor for lung complications in COVID-19 patients.

High sugar diets can increase susceptibility to bacterial infection in Drosophila melanogaster.

Overnutrition with dietary sugar can worsen infection outcomes in diverse organisms including insects and humans, through generally unknown mechanisms. In the present study, we show that adult Drosophila melanogaster fed high-sugar diets became more susceptible to infection by the Gram-negative bacteria Providencia rettgeri and Serratia marcescens. We found that P. rettgeri and S. marcescens proliferate more rapidly in D. melanogaster fed a high-sugar diet, resulting in increased probability of host death. D. melanogaster become hyperglycemic on the high-sugar diet, and we find evidence that the extra carbon availability may promote S. marcescens growth within the host. However, we found no evidence that increased carbon availability directly supports greater P. rettgeri growth. D. melanogaster on both diets fully induce transcription of antimicrobial peptide (AMP) genes in response to infection, but D. melanogaster provided with high-sugar diets show reduced production of AMP protein. Thus, overnutrition with dietary sugar may impair host immunity at the level of AMP translation. Our results demonstrate that dietary sugar can shape infection dynamics by impacting both host and pathogen, depending on the nutritional requirements of the pathogen and by altering the physiological capacity of the host to sustain an immune response.

Antimicrobial Protein LECT2-b Helps Maintain Gut Microbiota Homeostasis via Selectively Targeting Certain Pathogenic Bacteria.

Antimicrobial peptides/proteins (AMPs) constitute a critical component of gut immunity in animals, protecting the gut from pathogenic bacteria. However, the interactions between AMPs and gut microbiota remain elusive. In this study, we show that leukocyte-derived chemotaxin-2 (LECT2)-b, a recently discovered AMP, helps maintain gut homeostasis in grass carp (Ctenopharyngodon idella), one of the major farmed fish species globally, by directly regulating the gut microbiota. Knockdown of LECT2-b resulted in dysregulation of the gut microbiota. Specifically, LECT2-b deficiency led to the dominance of Proteobacteria, consisting of proinflammatory bacterial species, over Firmicutes, which includes anti-inflammatory bacteria. In addition, the opportunistic pathogenic bacteria genus Aeromonas became the dominant genus replacing the probiotic bacteria Lactobacillus and Bacillus. Further analysis revealed that this effect was due to the direct and selective inhibition of certain pathogenic bacterial species by LECT2-b. Moreover, LECT2-b knockdown promoted biofilm formation by gut microbiota, resulting in tissue damage and inflammation. Importantly, LECT2-b treatment alleviated the negative effects induced by LECT2-b knockdown. These findings highlight the crucial role of LECT2-b in maintaining the gut microbiota homeostasis and mucosal health. Overall, our study provides important data for understanding the roles of AMPs in the regulation of gut homeostasis in animals.

Process intensification for the production of a C-tagged antimicrobial peptide in Escherichia coli – First steps toward a platform technology

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

Expression of chemerin in intestinal mucosa of calves with comparable expression level with other antimicrobial proteins.

Neonatal calves are highly susceptible to infectious disorders including diarrhea. Therefore, epithelial innate immunity, including antimicrobial peptides/proteins (AMPs), is important during the early stage of their lives. Chemerin, a multifunctional protein that was originally identified as a chemokine, possesses a potent antimicrobial activity. The present study investigated the expression levels of chemerin in the gastrointestinal (GI) tract of growing calves. Chemerin and its coding gene, retinoic acid receptor responder protein 2 (RARRES2), were highly expressed in duodenum, jejunum, and ileum compared with other parts of the GI tract. Immunohistochemistry demonstrated that chemerin-producing cells were localized in the crypt of the intestinal mucosa. Finally, the expression level of RARRES2 was higher compared with those of other major AMPs in duodenum, although it was lower compared with that of enteric β-defensin but mostly higher than those of other AMPs in jejunum and ileum at various ages in calves. The expression levels of RARRES2 were not influenced by the age of calves in duodenum and jejunum, whereas a higher expression level of RARRES2 in ileum was observed in younger calves. This study revealed that chemerin is produced in the small intestine of calves and has the potential to contribute to the gut epithelial barrier system.

Crosstalk between keratinocytes and immune cells in inflammatory skin diseases

Cutaneous homeostasis is maintained by dynamic cellular communications between different cell types in the skin through interactions with various mediators, including cytokines, chemokines and antimicrobial peptides/proteins (AMPs). Keratinocytes, as the major cell type of the epidermis, not only form a passive physical barrier, but also actively participate in the pathogenesis of many, if not all, inflammatory skin diseases. Keratinocytes highly interact with immune cells to shape, amplify or regulate inflammatory responses, thus triggering and/or sustaining these inflammatory skin diseases. In this review, crosstalk between keratinocytes and immune cells is summarized, and its contributions to two major inflammatory skin disorders including psoriasis and atopic dermatitis are highlighted.

Cloning of selected body surface antimicrobial peptide/protein genes of Atlantic salmon and their responses to Aeromonas salmonicida

Antimicrobial peptides/proteins (AMPs), the important host-defence molecules with antimicrobial and immunomodulatory properties, effectively ward off pathogenic organisms. The responses of such molecules of Atlantic salmon during Aeromonas salmonicida infection that causes losses to salmon farms are not clearly described. Two trials were carried out to understand the regulation of two known AMP genes (cathelicidins, cath1, 2) and five newly cloned AMP genes (beta-defensins, defb1-4 and l-amino acid oxidase, lao); in vivo responses following intraperitoneal injection of A. salmonicida vaccine and in vitro responses upon exposure of gill tissues to A. salmonicida. The vaccine injection induced cath1, cath2 and lao expression in the gills at 24 h post injection. The live A. salmonicida exposure of Atlantic salmon gill tissue explant revealed the upregulation of the cath1 and cath2 within 48 h. The defb genes were not altered by either injection or exposure to A. salmonicida. Based on the evidence we suggest that the cathelicidins (cath1 and cath2) are A. salmonicida-inducible AMPs and could be possible biomarkers of infection.

Tissue-dependent induction of antimicrobial peptide genes after body wall injury in house fly (Musca domestica) larvae.

Injury of the insect body wall, which enables environmental microorganisms to invade into insect tissues, induces innate immune responses including the induction of antimicrobial peptides (AMPs) in flies and silkworms. Here, house fly (Musca domestica) larvae and pupae were injured using a needle and the effects on the expression of genes encoding AMPs were examined. The expression of AMP genes including defensin, attacin, diptericin, and sarcotoxin II dramatically increased in both larvae and pupae after injury of the body wall, indicating that innate immune responses were induced. Furthermore, the injury-dependent expression of AMP genes was examined in larval tissues including fat bodies, hemocytes, salivary glands, and digestive tracts. Injury-dependent AMP gene expression was observed in salivary glands, hemocytes, and fat bodies, but not in digestive tracts. The degree of the transcriptional induction of each gene differed among tissues, suggesting that their expression is governed by complex regulatory machinery and that AMPs have tissue-specific functions. To further examine the properties of the AMPs, we examined the antimicrobial activities of partial synthetic peptides corresponding to portions of the predicted AMP proteins deduced from the AMP genes. A synthetic peptide exhibited antimicrobial activity, indicating that these injury-inducible genes are potential medicinal resources.

De novo assembly and functional annotation of the emma field cricket (Teleogryllus emma) transcriptome

Abstract In recent years, insects have been used in animal-based foods for sustainable live-stock management and in the human diet as therapeutic foods. These insects are classified as edible and are widely consumed around the world for their broad medicinal and nutritional values. Antimicrobial peptides/proteins (AMPs) confer resistance to microbial pathogens and contribute to the immune function of insects. The emma field cricket (Teleogryllus emma) is widely consumed around the world for its broad-spectrum therapeutic effects. However, the absence of genetic information on T. emma is an obstacle to understanding the diversity and activity of its AMPs. Hence, deep sequencing of the T. emma transcriptome and expression profiling are essential. We report whole-body de novo transcriptome profiling of T. emma and analysis of its potential bacterial immunization-induced AMPs. We assembled 18.2 Gb of Illumina reads, generated 102,311 transcripts (minimum length: >500 bp, N50: 2585 bp), and identified 67,838 unigenes. Of these unigenes, 39,937 (58.87%) matched known proteins in a Basic Local Alignment Search Tool search of the National Center for Biotechnology Information database. Functional analysis demonstrated that the 39,937 unigenes clustered under 5707 Gene Ontology terms. In addition, we identified 10 cDNAs encoding new AMPs via an in-silico approach using public databases. Our transcriptome data set and AMP profile greatly improve the understanding of T. emma genetics and provide a huge number of gene sequences, including those for genes of known and unknown function, for further study.

The GCN2-ATF4 Signaling Pathway Induces 4E-BP to Bias Translation and Boost Antimicrobial Peptide Synthesis in Response to Bacterial Infection

Bacterial infection often leads to suppression of mRNA translation, but hosts are nonetheless able to express immune response genes through as yet unknown mechanisms. Here, we use a Drosophila model to demonstrate that antimicrobial peptide (AMP) production during infection is paradoxically stimulated by the inhibitor of cap-dependent translation, 4E-BP (eIF4E-binding protein; encoded by the Thor gene). We found that 4E-BP is induced upon infection with pathogenic bacteria by the stress-response transcription factor ATF4 and its upstream kinase, GCN2. Loss of gcn2, atf4, or 4e-bp compromised immunity. While AMP transcription is unaffected in 4e-bp mutants, AMP protein levels are substantially reduced. The 5' UTRs of AMPs score positive in cap-independent translation assays, andthis cap-independent activity is enhanced by 4E-BP. These results are corroborated invivo using transgenic 5' UTR reporters. These observations indicate that ATF4-induced 4e-bp contributes to innate immunity by biasing mRNA translation toward cap-independent mechanisms, thus enhancing AMP synthesis.

Friends or Foes? Host defense (antimicrobial) peptides and proteins in human skin diseases.

Host defense peptides/proteins (HDPs), also known as antimicrobial peptides/proteins (AMPs), are key molecules in the cutaneous innate immune system. AMPs/HDPs historically exhibit broad-spectrum killing activity against bacteria, enveloped viruses, fungi and several parasites. Recently, AMPs/HDPs were shown to have important biological functions, including inducing cell proliferation, migration and differentiation; regulating inflammatory responses; controlling the production of various cytokines/chemokines; promoting wound healing; and improving skin barrier function. Despite the fact that AMPs/HDPs protect our body, several studies have hypothesized that these molecules actively contribute to the pathogenesis of various skin diseases. For example, AMPs/HDPs play crucial roles in the pathological processes of psoriasis, atopic dermatitis, rosacea, acne vulgaris, systemic lupus erythematosus and systemic sclerosis. Thus, AMPs/HDPs may be a double-edged sword, promoting cutaneous immunity while simultaneously initiating the pathogenesis of some skin disorders. This review will describe the most common skin-derived AMPs/HDPs (defensins, cathelicidins, S100 proteins, ribonucleases and dermcidin) and discuss the biology and both the positive and negative aspects of these AMPs/HDPs in skin inflammatory/infectious diseases. Understanding the regulation, functions and mechanisms of AMPs/HDPs may offer new therapeutic opportunities in the treatment of various skin disorders.

Transcriptome Profiling and In Silico Analysis of the Antimicrobial Peptides of the Grasshopper Oxya chinensis sinuosa.

Antimicrobial peptides/proteins (AMPs) are present in all types of organisms, from microbes and plants to vertebrates and invertebrates such as insects. The grasshopper Oxya chinensis sinuosa is an insect species that is widely consumed around the world for its broad medicinal value. However, the lack of available genetic information for this species is an obstacle to understanding the full potential of its AMPs. Analysis of the O. chinensis sinuosa transcriptome and expression profile is essential for extending the available genetic information resources. In this study, we determined the whole-body transcriptome of O. chinensis sinuosa and analyzed the potential AMPs induced by bacterial immunization. A high-throughput RNA-Seq approach generated 94,348 contigs and 66,555 unigenes. Of these unigenes, 36,032 (54.14%) matched known proteins in the NCBI database in a BLAST search. Functional analysis demonstrated that 38,219 unigenes were clustered into 5,499 gene ontology terms. In addition, 26 cDNAs encoding novel AMPs were identified by an in silico approach using public databases. Our transcriptome dataset and AMP profile greatly improve our understanding of O. chinensis sinuosa genetics and provide a huge number of gene sequences for further study, including genes of known importance and genes of unknown function.

Insect antimicrobial peptides: potential tools for the prevention of skin cancer.

Antimicrobial peptides/proteins (AMPs) are biologically active molecules with diverse structural properties that are produced by mammals, plants, insects, ticks, and microorganisms. They have a range of antibacterial, antifungal, antiviral, and even anticancer activities, and their biological properties could therefore be exploited for therapeutic and prophylactic applications. Cancer and cancer drug resistance are significant current health challenges, so the development of innovative cancer drugs with minimal toxicity toward normal cells and novel modes of action that can evade resistance may provide a new direction for anticancer therapy. The skin is the first line of defense against heat, sunlight, injury, and infection, and skin cancer is thus the most common type of cancer. The skin that has been exposed to sunlight is particularly susceptible, but lesions can occur anywhere on the body. Skin cancer awareness and self-efficacy are necessary to improve sun protection behavior, but more effective preventative approaches are also required. AMPs may offer a new prophylactic approach against skin cancer. In this mini review, we draw attention to the potential use of insect AMPs for the prevention and treatment of skin cancer.

Antimicrobial Peptide LL-37 Is Both a Substrate of Cathepsins S and K and a Selective Inhibitor of Cathepsin L.

Lung cysteine cathepsins B, K, L, and S contribute to physiological and pathological processes including degradation of antimicrobial peptides/proteins (AMPs) such as surfactant protein SP-A, lactoferrin, secretory leukocyte peptidase inhibitor, and beta-defensins-2 and -3. Substantial amounts of uncleaved LL-37, a 37-mer cationic AMP, were observed in the sputum of patients with cystic fibrosis (CF). Nevertheless LL-37 was degraded after prolonged incubation in CF sputum, and the hydrolysis was blocked by E-64, a selective inhibitor of cysteine proteases. Cathepsins K and S, expressed in human alveolar macrophages, thoroughly hydrolyzed LL-37 in vitro, whereas it competitively inhibited cathepsin L (Ki = 150 nM). Cleavage of LL-37 by cathepsins S and K impaired its antimicrobial activity against Pseudomonas aeruginosa and Staphylococcus aureus, in a time- and concentration-dependent manner. The exchange of residues 67 and 205 in the S2 pockets of cathepsins L (Leu67Tyr/Ala205Leu) and K (Tyr67Leu/Leu205Ala) switched the specificity of these mutants toward LL-37. Molecular modeling suggested that LL-37 interacted with the active site of cathepsin L in both forward (i.e., substrate-like) and reverse orientations with similar binding energies. Our data support the hypothesis that cysteine cathepsins modulate the innate immunity response by degrading distinct and representative members of the AMP family.

Insect antimicrobial peptides and their applications.

Insects are one of the major sources of antimicrobial peptides/proteins (AMPs). Since observation of antimicrobial activity in the hemolymph of pupae from the giant silk moths Samia Cynthia and Hyalophora cecropia in 1974 and purification of first insect AMP (cecropin) from H. cecropia pupae in 1980, over 150 insect AMPs have been purified or identified. Most insect AMPs are small and cationic, and they show activities against bacteria and/or fungi, as well as some parasites and viruses. Insect AMPs can be classified into four families based on their structures or unique sequences: the α-helical peptides (cecropin and moricin), cysteine-rich peptides (insect defensin and drosomycin), proline-rich peptides (apidaecin, drosocin, and lebocin), and glycine-rich peptides/proteins (attacin and gloverin). Among insect AMPs, defensins, cecropins, proline-rich peptides, and attacins are common, while gloverins and moricins have been identified only in Lepidoptera. Most active AMPs are small peptides of 20-50 residues, which are generated from larger inactive precursor proteins or pro-proteins, but gloverins (~14kDa) and attacins (~20kDa) are large antimicrobial proteins. In this mini-review, we will discuss current knowledge and recent progress in several classes of insect AMPs, including insect defensins, cecropins, attacins, lebocins and other proline-rich peptides, gloverins, and moricins, with a focus on structural-functional relationships and their potential applications.

Pseudomonas aeruginosa in Musca domestica L.: Temporospatial Examination of Bacteria Population Dynamics and House Fly Antimicrobial Responses

House flies associate with microbes throughout their life history. Bacteria ingested by adult flies enter the alimentary canal and face a hostile environment including antimicrobial defenses. Because the outcome of this interaction impacts bacterial survival and dissemination, our primary objective was to understand the temporospatial dynamics of fly-bacteria associations. We concurrently examined the temporospatial fate of GFP-expressing Pseudomonas aeruginosa (GFP-P. aeruginosa) in the house fly alimentary canal along with antimicrobial peptide (AMP) expression. Motile, viable GFP-P. aeruginosa were found in all regions of the alimentary canal and were culturable throughout the observation period (2–24 h). A significant decrease in recoverable bacteria occurred between 2 and12 h, followed by an increase between 12 and 24 h. qRT-PCR analysis showed expression of the AMPs cecropin, diptericin, and defensin both locally (gut) and systemically. Furthermore, mRNA of all AMPs were expressed throughout gut tissues, with some tissue-specific temporal variation. Interestingly, fluctuation in recoverable P. aeruginosa was associated with AMP protein expression in the gut (immunofluorescent signal detection), but not with mRNA (qRTPCR). In regards to vector competence, flies excreted GFP-P. aeruginosa throughout the 24 h period, serving as both reservoirs and disseminators of this bacterium. Collectively, our data show flies can harbor and disseminate P. aeruginosa, and that the interactions of fly defenses with bacteria can influence vector competence.

Identification and Expression Profile Analysis of Antimicrobial Peptide/Protein in Asian Corn Borer, Ostrinia furnacalis (Guenée)

Antimicrobial peptides/proteins (AMPs) are a group of immune proteins that exhibit strong antibiotic properties against numerous infectious bacterial strains. They are evolutionarily conserved and present in every kingdom and phylum, ranging from prokaryotes to humans. We analyzed the transciptome from the larvae of Asian corn borer, Ostrinia furnacalis (Guenée), and identified several putative AMP transcripts, OfgLys5, OfgLys6, OfgLys10, OfgAtt, and OfgIID. OfgLys5, OfgLys6, and OfgLys10 are all highly homologous with c-type lysozymes, and OfgAtt shows significant identities with Lepidoptera attacin. The amino acid sequence of OfgLys5 and OfgLys6 possessed all conserved features critical for fundamental structure and function of c-type lysozyme, including the two catalytic sites, Glu32 and Asp50. OfgAtt is a typical glycine-rich protein. The antimicrobial activity of O. furnacalis hemolymph increased significantly after injection with Escherichia coli, Micrococcus luteus, or Beauveria bassiana. OfgAtt, IDD, and Lys6 are expressed at low level prior to the challenge, but strongly induced against Gram-positive and negative bacteria, and fungi. Under the same inducement conditions, the transcripts of these three genes elevated most when fifth instar larvae were injected. Therefore, O. furnacalis larvae are induced to produce antimicrobial materials in the hemolymph after the infection, and increase of lysozyme and attacin may contribute to the antimicrobial activity.

Immune stimulant effects of a nucleotide-rich baker's yeast extract in the kuruma shrimp, Marsupenaeus japonicus

Immune stimulant effects of a nucleotide-rich baker's yeast extract (Vertex IG20) were investigated in the kuruma shrimp, Marsupenaeus japonicus by examining expression of anti-microbial peptides/proteins (AMPs) such as penaeidin (MjPen), crustin (MjCrus) and lysozyme (MjLyz) genes. Furthermore, to confirm that the baker's yeast extract-induced AMPs were functional, we also assessed the effect of its oral administration on resistance to Vibrio nigripulchritudo infection in the kuruma shrimp. Our results demonstrate that baker's yeast extract-injected and fed shrimps displayed a significant up-regulation of MjPen, MjCrus and MjLyz gene expression in the lymphoid organ. Moreover, significantly increased (P<0.01) resistance to the bacterial pathogen in term of better post infection survival (66.6%) was observed in the shrimp fed with the yeast extract-incorporated diet compared with the control diet fed group (8.3%). The present study indicates the immunostimulatory effects of the nucleotide-rich baker's yeast extract on the kuruma shrimp immune system and supports its potential use in aquaculture.

Outer Membrane Lipoprotein Lpp Is Gram-negative Bacterial Cell Surface Receptor for Cationic Antimicrobial Peptides

Antimicrobial peptides/proteins (AMPs) are important components of the host innate defense mechanisms. Here we demonstrate that the outer membrane lipoprotein, Lpp, of Enterobacteriaceae interacts with and promotes susceptibility to the bactericidal activities of AMPs. The oligomeric Lpp was specifically recognized by several cationic α-helical AMPs, including SMAP-29, CAP-18, and LL-37; AMP-mediated bactericidal activities were blocked by anti-Lpp antibody blocking. Blebbing of the outer membrane and increase in membrane permeability occurred in association with the coordinate internalization of Lpp and AMP. Interestingly, the specific binding of AMP to Lpp was resistant to divalent cations and salts, which were able to inhibit the bactericidal activities of some AMPs. Furthermore, using His-tagged Lpp as a ligand, we retrieved several characterized AMPs, including SMAP-29 and hRNase 7, from a peptide library containing crude mammalian cell lysates. Overall, this study explores a new mechanism and target of antimicrobial activity and provides a novel method for screening of antimicrobials for use against drug-resistant bacteria.

The Antimicrobial Defense of the Pacific Oyster, Crassostrea gigas. How Diversity may Compensate for Scarcity in the Regulation of Resident/Pathogenic Microflora

Healthy oysters are inhabited by abundant microbial communities that vary with environmental conditions and coexist with immunocompetent cells in the circulatory system. In Crassostrea gigas oysters, the antimicrobial response, which is believed to control pathogens and commensals, relies on potent oxygen-dependent reactions and on antimicrobial peptides/proteins (AMPs) produced at low concentrations by epithelial cells and/or circulating hemocytes. In non-diseased oysters, hemocytes express basal levels of defensins (Cg-Defs) and proline-rich peptides (Cg-Prps). When the bacterial load dramatically increases in oyster tissues, both AMP families are driven to sites of infection by major hemocyte movements, together with bactericidal permeability/increasing proteins (Cg-BPIs) and given forms of big defensins (Cg-BigDef), whose expression in hemocytes is induced by infection. Co-localization of AMPs at sites of infection could be determinant in limiting invasion as synergies take place between peptide families, a phenomenon which is potentiated by the considerable diversity of AMP sequences. Besides, diversity occurs at the level of oyster AMP mechanisms of action, which range from membrane lysis for Cg-BPI to inhibition of metabolic pathways for Cg-Defs. The combination of such different mechanisms of action may account for the synergistic activities observed and compensate for the low peptide concentrations in C. gigas cells and tissues. To overcome the oyster antimicrobial response, oyster pathogens have developed subtle mechanisms of resistance and evasion. Thus, some Vibrio strains pathogenic for oysters are equipped with AMP-sensing systems that trigger resistance. More generally, the known oyster pathogenic vibrios have evolved strategies to evade intracellular killing through phagocytosis and the associated oxidative burst.