Direct Antimicrobial Properties Research Articles

Progress of Antimicrobial Mechanisms of Stilbenoids.

Antimicrobial drugs have made outstanding contributions to the treatment of pathogenic infections. However, the emergence of drug resistance continues to be a major threat to human health in recent years, and therefore, the search for novel antimicrobial drugs is particularly urgent. With a deeper understanding of microbial habits and drug resistance mechanisms, various creative strategies for the development of novel antibiotics have been proposed. Stilbenoids, characterized by a C6-C2-C6 carbon skeleton, have recently been widely recognized for their flexible antimicrobial roles. Here, we comprehensively summarize the mode of action of stilbenoids from the viewpoint of their direct antimicrobial properties, antibiofilm and antivirulence activities and their role in reversing drug resistance. This review will provide an important reference for the future development and research into the mechanisms of stilbenoids as antimicrobial agents.

The Role of Neutrophil Extracellular Traps in the Outcome of Malignant Epitheliomas: Significance of CA215 Involvement

Neutrophil extracellular traps (NETs) were originally discovered as a part of the innate immune response of the host to bacteria. They form a web-like structure that can immobilize microorganisms or exhibit direct antimicrobial properties, such as releasing reactive oxygen species (ROS). NETs are established when neutrophils undergo a sort of cellular death following exposure to ROS, chemokines, cytokines, or other soluble factors. This process results in the release of the neutrophil’s DNA in a web-like form, which is decorated with citrullinated histones (H3/H4-cit), neutrophil elastase (NE), and myeloperoxidase (MPO). Emerging studies have put into perspective that NETs play an important role in oncology as they were shown to influence tumor growth, malignant initiation, and proliferation, mediate the transition from endothelial to mesenchymal tissue, stimulate angiogenesis or metastasis, and can even help cancer cells evade the immune response. The role of NETs in cancer therapy resides in their ability to form and act as a mechanical barrier that will provide the primary tumor with a reduced response to irradiation or pharmaceutical penetration. Subsequently, cancer cells are shown to internalize NETs and use them as a strong antioxidant when pharmaceutical treatment is administered. In this review, we explored the role of NETs as part of the tumor microenvironment (TME), in the context of malignant epitheliomas, which are capable of an autonomous production of CA215, a subvariant of IgG, and part of the carcinoembryonic antigen (CEA) superfamily. Studies have shown that CA215 has a functional Fc subdivision able to activate the Fc-gamma-RS receptor on the surface of neutrophils. This activation may afterward stimulate the production of NETs, thus indicating CA215 as a potential factor in cancer therapy surveillance.

New Evidence for the Role of Pituitary Adenylate Cyclase-Activating Polypeptide as an Antimicrobial Peptide in Teleost Fish.

Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) is a multifunctional neuropeptide that is widely distributed and conserved across species. We have previously shown that in teleost fish, PACAP not only possesses direct antimicrobial properties but also immunomodulatory effects against the bacterial pathogens Flavobacterium psychrophilum and Pseudomonas aeruginosa using in vitro and in vivo experiments. These previous results suggest PACAP can be used as an alternative to antibiotics to prevent and/or treat bacterial infections in the aquaculture industry. To accomplish this goal, more studies are needed to better understand the effect of PACAP on pathogens affecting fish in live infections. In the present study, the transcripts PACAP, PRP/PACAP, and VPAC2 receptor were examined in rainbow trout (Oncorhynchus mykiss) naturally infected with Yersinia ruckeri, which exhibited an increase in their expression in the spleen when compared to healthy fish. Synthetic Clarias gariepinus PACAP-38 has direct antimicrobial activity on Y. ruckeri and inhibits up to 60% of the bacterial growth when the peptide is at concentrations between 50 and 100 µM in TSB. The growth inhibition increased up to 90% in the presence of 12.5 µM of PACAP-38 when salt-free LB broth was used instead of TSB. It was also found to inhibit Y. ruckeri growth in a dose-dependent manner when the rainbow trout monocyte/macrophage-like cell line (RTS11) was pre-treated with lower concentrations of the peptide (0.02 and 0.1 µM) before going through infection. Differential gene expression was analyzed in this in vitro model. Overall, the results revealed new evidence to support the role of PACAP as an antimicrobial and immunomodulatory peptide treatment in teleosts.

Recent advances and prospects in purification and heterologous expression of lactoferrin

AbstractLactoferrin is an 80 kDa iron‐binding glycoprotein of the transferrin family, which is abundant in milk and in most biological fluids. Besides the direct antimicrobial properties, lactoferrin can regulate immune responses and prevent infection and septic shock. Membrane adsorption and chromatographic methods are usually used for separating and purifying of lactoferrin from bovine milk. However, the content of lactoferrin in milk is relatively low, resulting in a high cost of producing lactoferrin. Meanwhile, as a heterologous protein applied to the human body, it may cause a certain antigen response. Therefore, obtaining a large amount of biologically active lactoferrin has always been a research hot topic. This review first introduced the physical and chemical properties, and biological activities of lactoferrin. Then, the current recombinant expression systems for lactoferrin were summarized. Finally, the current challenges and the future development trends of efficient synthesis of lactoferrin were discussed.

Linear-Chain Nanostructured Carbon with a Silver Film Plated on Metal Components Has a Promising Effect for the Treatment of Periprosthetic Joint Infection

Background: Due to the aging of the world population, the number of joint diseases, along with the number of arthroplasties, has increased, simultaneously increasing the amount of complications, including periprosthetic joint infection (PPI). In this study, to combat a PPI, we investigated the antimicrobial properties of the new composite cover for titanium implants, silver-doped carbyne-like carbon (S-CLC) film. Methods: The first assay investigated the antimicrobial activity against Pseudomonas aeruginosa and releasing of silver ions from S-CLC films into growth media covered with S-CLC with a thickness of 1, 2, and 4 mm. The second assay determined the direct antibacterial properties of the S-CLC film’s surface against Staphylococcus aureus, Enterococcus faecalis, or P. aeruginosa. The third assay studied the formation of microbial biofilms of S. aureus or P. aeruginosa on the S-CLC coating. Silver-doped carbyne-like carbon (S-CLC)-covered or titanium plates alone were used as controls. Results: S-CLC films, compared to controls, prevented P. aeruginosa growth on 1 mm thickness agar; had direct antimicrobial properties against S. aureus, E. faecalis, and P. aeruginosa; and could prevent P. aeruginosa biofilm formation. Conclusions: S-CLC films on the Ti surface could successfully fight the most common infectious agent in PPI, and prevented biofilm formation.

Insect Defense Proteins and Peptides.

The composition of insect hemolymph can change depending on many factors, e.g. access to nutrients, stress conditions, and current needs of the insect. In this chapter, insect immune-related polypeptides, which can be permanently or occasionally present in the hemolymph, are described. Their division into peptides or low-molecular weight proteins is not always determined by the length or secondary structure of a given molecule but also depends on the mode of action in insect immunity and, therefore, it is rather arbitrary. Antimicrobial peptides (AMPs) with their role in immunity, modes of action, and classification are presented in the chapter, followed by a short description of some examples: cecropins, moricins, defensins, proline- and glycine-rich peptides. Further, we will describe selected immune-related proteins that may participate in immune recognition, may possess direct antimicrobial properties, or can be involved in the modulation of insect immunity by both abiotic and biotic factors. We briefly cover Fibrinogen-Related Proteins (FREPs), Down Syndrome Cell Adhesion Molecules (Dscam), Hemolin, Lipophorins, Lysozyme, Insect Metalloproteinase Inhibitor (IMPI), and Heat Shock Proteins. The reader will obtain a partial picture presenting molecules participating in one of the most efficient immune strategies found in the animal world, which allow insects to inhabit all ecological land niches in the world.

Heterologous Expression and Purification of the Antimicrobial Peptide Buforin II

Abstract Antimicrobial peptides are natural substances that have played a role in the development of the adaptive immune system, and are currently involved in the prevention of infections, through their direct antimicrobial and immunomodulatory properties. While the amino acid composition and spatial structure vary, most antibacterial peptides have a positive surface charge, which allows them to bind to the negative bacterial membranes. Buforin II is a widely studied antimicrobial peptide first obtained through the structural modification of buforin I, a peptide isolated from Bufo gargarizans. The peptide showed significant antibacterial activity against Gram-positive and Gram-negative bacterial strains. The mechanism of action of buforin II differs from that of other antimicrobial peptides, as it binds directly to bacterial DNA and RNA. The aim of our study was to obtain recombinant buforin II with a ubiquitin fusion partner, through heterologous expression in Escherichia coli Rosetta™ (DE3)pLysS cells, using a laboratory scale bioreactor. The incubation of expression host cells in a bioreactor allowed the constant monitoring and control of the process parameters, leading to high biomass levels and an increased production rate of the peptide. The parameters used during incubation were: 37°C, pH=6.9 and dissolved oxygen level above 40%. Purification of the recombinant protein was accomplished by affinity chromatography using a Ni-chelate solid phase to which the 10xHistag of our construct showed affinity. Method optimisation consisted in the use of gradient and linear elution, of which the latter was found to be more effective. Digestion of the fusion partner from the target peptide was performed with ubiquitin carboxyl-terminal hydrolase enzyme. The expression and purification protocols developed in our experiment allow the production of a significant amount of buforin II, allowing its use for further research. Furthermore, the presented methods could be suitable for industrial production of the recombinant peptide..

Nicotinamide Limits Replication of Mycobacterium tuberculosis and Bacille Calmette-Guérin Within Macrophages.

Novel antimicrobials for treatment of Mycobacterium tuberculosis are needed. We hypothesized that nicotinamide (NAM) and nicotinic acid (NA) modulate macrophage function to restrict M. tuberculosis replication in addition to their direct antimicrobial properties. Both compounds had modest activity in 7H9 broth, but only NAM inhibited replication in macrophages. Surprisingly, in macrophages NAM and the related compound pyrazinamide restricted growth of bacille Calmette-Guérin but not wild-type Mycobacterium bovis, which both lack a functional nicotinamidase/pyrazinamidase (PncA) rendering each strain resistant to these drugs in broth culture. Interestingly, NAM was not active in macrophages infected with a virulent M. tuberculosis mutant encoding a deletion in pncA. We conclude that the differential activity of NAM and nicotinic acid on infected macrophages suggests host-specific NAM targets rather than PncA-dependent direct antimicrobial properties. These activities are sufficient to restrict attenuated BCG, but not virulent wild-type M. bovis or M. tuberculosis.

Human umbilical-cord mesenchymal stem cells inhibit bacterial growth and alleviate antibiotic resistance in neonatal imipenem-resistant Pseudomonas aeruginosa infection.

Human umbilical-cord mesenchymal stem cells (hUCMSCs) are a safe and convenient source of MSCs and have shown beneficial effects in neonatal infection and sepsis animal models. However, the factors leading to improved outcomes are still unclear. The aim of this study was to investigate the antibacterial effect and regulation of antimicrobial resistance of hUCMSCs. We separated imipenem-resistant Pseudomonas aeruginosa (PA) from neonates and incubated it with hUCMSCs as well as their culture medium. Assessment of direct inhibition of bacterial growth was done by counting CFUs. The concentration of antibacterial peptides in the culture medium of hUCMSCs was measured. Standard PA was inoculated with a sub-inhibitory concentration of imipenem with and without hUCMSC conditioned medium and antimicrobial peptides. The sensitivity to imipenem was detected until PA showed resistance to imipenem. Outer membrane protein (OprD2) mRNA expression in PA before and after the induction of imipenem resistance was analysed. We found that HUCMSCs possessed direct antimicrobial properties against bacteria and could alleviate antibiotic resistance via reserving OprD2 expression in PA.

Ibuprofen lacks direct antimicrobial properties for the treatment of urinary tract infection isolates.

The high incidence of urinary tract infection (UTI) among women and children, in combination with a lack of antibiotic efficacy with regard to pathogen eradication and recurrence prevention, as well as the negative side effects associated with antibiotics, has led researchers to explore the role of non-steroidal anti-inflammatory drugs as a primary management strategy. The aim of this study was to determine whether ibuprofen (IBU) or one of its major metabolites, 2-carboxyibuprofen (CIBU), could affect the growth and adhesion of the two most common uropathogens, Escherichia coli and Enterococcus faecalis. The bacterial growth and adhesion to the urothelial cells of E. coli UTI89 and E. faecalis 1131 in the presence of physiologically relevant concentrations of IBU and CIBU were assessed. The effect of IBU on bacterial adhesion to urothelial cells was also assessed following exposure to trimethoprim/sulfamethoxazole (TMP/SMX) and nitrofurantoin. Bacterial growth was not affected by IBU. Further, only at high levels of IBU not regularly found in the bladder was there a significant increase in E. faecalis 1131 attachment at growth inhibitory concentrations of TMP/SMX. There was no effect on the attachment of E. faecalis or E. coli to urothelial cells in the presence of nitrofurantoin. These studies indicate that the beneficial effects of IBU for UTI management are likely mediated through its anti-inflammatory properties rather than direct interactions with uropathogens in the bladder.

Effects of human antimicrobial cryptides identified in apolipoprotein B depend on specific features of bacterial strains

Cationic Host Defense Peptides (HDPs) are endowed with a broad variety of activities, including direct antimicrobial properties and modulatory roles in the innate immune response. Even if it has been widely demonstrated that bacterial membrane represents the main target of peptide antimicrobial activity, the molecular mechanisms underlying membrane perturbation by HDPs have not been fully clarified yet. Recently, two cryptic HDPs have been identified in human apolipoprotein B and found to be endowed with a broad-spectrum antimicrobial activity, and with anti-biofilm, wound healing and immunomodulatory properties. Moreover, ApoB derived HDPs are able to synergistically act in combination with conventional antibiotics, while being not toxic for eukaryotic cells. Here, by using a multidisciplinary approach, including time killing curves, Zeta potential measurements, membrane permeabilization assays, electron microscopy analyses, and isothermal titration calorimetry studies, the antimicrobial effects of ApoB cryptides have been analysed on bacterial strains either susceptible or resistant to peptide toxicity. Intriguingly, it emerged that even if electrostatic interactions between negatively charged bacterial membranes and positively charged HDPs play a key role in mediating peptide toxicity, they are strongly influenced by the composition of negatively charged bacterial surfaces and by defined extracellular microenvironments.

IL-26, a Cytokine With Roles in Extracellular DNA-Induced Inflammation and Microbial Defense.

Interleukin 26 (IL-26) is the most recently identified member of the IL-20 cytokine subfamily, and is a novel mediator of inflammation overexpressed in activated or transformed T cells. Novel properties have recently been assigned to IL-26, owing to its non-conventional cationic, and amphipathic features. IL-26 binds to DNA released from damaged cells and, as a carrier molecule for extracellular DNA, links DNA to inflammation. This observation suggests that IL-26 may act both as a driver and an effector of inflammation, leading to the establishment of a deleterious amplification loop and, ultimately, sustained inflammation. Thus, IL-26 emerges as an important mediator in local immunity/inflammation. The dysregulated expression and extracellular DNA carrier capacity of IL-26 may have profound consequences for the chronicity of inflammation. IL-26 also exhibits direct antimicrobial properties. This review summarizes recent advances on the biology of IL-26 and discusses its roles as a novel kinocidin.

Comparison of Antibacterial and Immunological Properties of Mesenchymal Stem/Stromal Cells from Equine Bone Marrow, Endometrium, and Adipose Tissue.

Equine mesenchymal stem/stromal cells (MSCs) are multipotent cells that are widely used for treatment of musculoskeletal injuries, and there is significant interest in expanding their application to nonorthopedic conditions. MSCs possess antibacterial and immunomodulatory properties that may be relevant for combating infection; however, comparative studies using MSCs from different origins have not been carried out in the horse, and this was the focus of this study. Our results showed that MSC-conditioned media attenuated the growth of Escherichia coli, and that this effect was, on average, more pronounced for endometrium (EM)-derived and adipose tissue (AT)-derived MSCs than for bone marrow (BM)-derived MSCs. In addition, the antimicrobial lipocalin-2 was expressed at mean higher levels in EM-MSCs than in AT-MSCs and BM-MSCs, and the bacterial component lipopolysaccharide (LPS) stimulated its production by all three MSC types. We also showed that MSCs express interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein-1, chemokine ligand-5, and Toll-like receptor 4, and that, in general, these cytokines were induced in all cell types by LPS. Low expression levels of the macrophage marker colony-stimulating factor 1 receptor were detected in BM-MSCs and EM-MSCs but not in AT-MSCs. Altogether, these findings suggest that equine MSCs from EM, AT, and BM have both direct and indirect antimicrobial properties that may vary between MSCs from different origins and could be exploited toward improvement of regenerative therapies for horses.

Rhamnolipids From Pseudomonas aeruginosa Are Elicitors Triggering Brassica napus Protection Against Botrytis cinerea Without Physiological Disorders.

Rhamnolipids (RLs) are amphiphilic molecules naturally produced by some bacteria with a large range of biological activities. Although some studies report their potential interest in plant protection, evaluation of their effects and efficiency on annual crops of worldwide agronomic interest is lacking. The main objective of this work was to investigate their elicitor and protective activities on rapeseed crop species while evaluating their physiological effects. Here we report that RLs from Pseudomonas aeruginosa secretome trigger an effective protection of Brassica napus foliar tissues toward the fungus Botrytis cinerea involving the combination of plant defense activation and direct antimicrobial properties. We demonstrated their ability to activate canonical B. napus defense responses including reactive oxygen species production, expression of defense genes, along with callose deposits and stomatal closure as efficient physical protections. In addition, microscopic cell death observations and electrolyte leakage measurements indicated that RLs trigger a hypersensitive response-like defense in this plant. We also showed that foliar spray applications of RLs do not induce deleterious physiological consequences on plant growth or chlorophyll content and that RL protective properties are efficient on several grown cultivars of rapeseed. To our knowledge, this is the first report of RLs as an elicitor that suppresses fungal disease on tissues of an annual crop species under greenhouse conditions. Our results highlight the dual mode of action of these molecules exhibiting plant protection activation and antifungal activities and demonstrate their potential for crop cultures as environmental-friendly biocontrol solution.

Amino Acid Block Copolymers with Broad Antimicrobial Activity and Barrier Properties.

Antimicrobial properties of a long-chain, synthetic, cationic, and hydrophobic amino acid block copolymer are reported. In 5 and 60 min time-kill assays, solutions of K100 L40 block copolymers (poly(l-lysine·hydrochloride)100 -b-poly(l-leucine)40 ) at concentrations of 10-100 µg mL-1 show multi-log reductions in colony forming units of Gram-positive and Gram-negative bacteria, as well as yeast, including multidrug-resistant strains. Driven by association of hydrophobic segments, K100 L40 copolymers form viscous solutions and self-supporting hydrogels in water at concentrations of 1 and 2 wt%, respectively. These K100 L40 preparations provide an effective barrier to microbial contamination of wounds, as measured by multi-log decreases of tissue-associated bacteria with deliberate inoculation of porcine skin explants, porcine open wounds, and rodent closed wounds with foreign body. Based on these findings, amino acid copolymers with the features of K100 L40 can combine potent, direct antimicrobial activity and barrier properties in one biopolymer for a new approach to prevention of wound infections.

Inflammatory Contribution of Platelets Revisited: New Players in the Arena of Inflammation.

Platelet-derived mediators, either in an autocrine or paracrine mode of action, regulate systemic and vascular inflammation as well as contribute to innate immune defense and also to regenerative mechanisms. This review reevaluates the impact of inflammatory mediators such as CXC-chemokine-Ligands, their receptors in modulating platelet functions and platelet survival, thereby influencing inflammatory or regenerative processes. We further explore the contribution of cyclophilin A and C-reactive protein in regulating thrombotic and hemostatic attributes of platelets. Moreover, we emphasize on the role of platelets as active components bridging the innate and adaptive immune responses, toll-like receptors on platelets, platelet interactions with the complement system, and platelet-derived thrombocidins exhibiting direct antimicrobial properties. As highlighted in this review, the multifaceted aspects of platelets and platelet-derived factors encourage further investigations in this intriguing and expansive but largely uncharted area of research in platelet biology.

Optimizing Crops for Biocontrol of Pests and Disease.

Volatile compounds and extrafloral nectar are common defenses of wild plants; however, in crops they bear an as-yet underused potential for biological control of pests and diseases. Odor emission and nectar secretion are multigene traits in wild plants, and thus form difficult targets for breeding. Furthermore, domestication has changed the capacity of crops to express these traits. We propose that breeding crops for an enhanced capacity for tritrophic interactions and volatile-mediated direct resistance to herbivores and pathogens can contribute to environmentally-friendly and sustainable agriculture. Natural plant volatiles with antifungal or repellent properties can serve as direct resistance agents. In addition, volatiles mediating tritrophic interactions can be combined with nectar-based food rewards for carnivores to boost indirect plant defense.

Regulation of the NADPH Oxidase RBOHD During Plant Immunity.

Pathogen recognition induces the production of reactive oxygen species (ROS) by NADPH oxidases in both plants and animals. ROS have direct antimicrobial properties, but also serve as signaling molecules to activate further immune outputs. However, ROS production has to be tightly controlled to avoid detrimental effects on host cells, but yet must be produced in the right amount, at the right place and at the right time upon pathogen perception. Plant NADPH oxidases belong to the respiratory burst oxidase homolog (RBOH) family, which contains 10 members in the model plant Arabidopsis thaliana. The perception of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs) leads to a rapid, specific and strong production of ROS, which is dependent on RBOHD. RBOHD is mainly controlled by Ca(2+) via direct binding to EF-hand motifs and phosphorylation by Ca(2+)-dependent protein kinases. Recent studies have, however, revealed a critical role for a Ca(2+)-independent regulation of RBOHD. The plasma membrane-associated cytoplasmic kinase BIK1 (BOTRYTIS-INDUCED KINASE1), which is a direct substrate of the PRR complex, directly interacts with and phosphorylates RBOHD upon PAMP perception. Impairment of these phosphorylation events completely abolishes the function of RBOHD in immunity. These results suggest that RBOHD activity is tightly controlled by multilayered regulations. In this review, we summarize recent advances in our understanding of the regulatory mechanisms controlling RBOHD activation.

Two thiadiazole compounds promote rice defence against Xanthomonas oryzae pv. oryzae by suppressing the bacterium's production of extracellular polysaccharides.

Thiazole, isothiazole, thiadiazole, and their derivatives are used to control various human, animal and plant diseases. In addition to having direct anti-microbial and anti-fungal properties, these compounds are thought to induce host defences, but the mechanism of defence induction remains poorly understood. This article reports that the thiadiazoles of zinc thiazole and bismerthiazol induce H2 O2 accumulation, up-regulation of defence-related genes, callose deposition and hypersensitive response-like cell death in rice leaves infected with Xanthomonas oryaze pv. oryzae (Xoo) strain ZJ173, but not in non-infected leaves. These defence responses in Xoo-infected leaves were suppressed by the exogenous application of catalase, which reduces H2 O2 accumulation. The application of extracellular polysaccharides (EPSs) extracted from strain ZJ173 significantly compromised rice defence against ZJ173 with or without thiadiazole treatment. The EPS-deficient Xoo mutant ∆gumH triggered a stronger defence than its parent strain ZJ173. The thiadiazole treatments reduced EPS production by strain ZJ173, but not by the thiadiazole-resistant strain 2-1-1, which is thiadiazole resistant in vivo, but not in vitro; moreover, enhanced defence was not detected in thiadiazole-treated rice inoculated with 2-1-1. Based on these data, we infer that zinc thiazole and bismerthiazol promote rice defence against Xoo by inhibiting the production of bacterial EPS.

Synthesis and the antiresistant activity of acylated and supramolecular derivatives of ellagic acid

The aim of our research was to obtain new, more soluble and biologically active derivatives of ellagic acid through acylation of phenol hydroxyls and obtaining supramolecular complexes with cyclodextrins, as well as to study the ability of the derivatives synthesized to eliminate the microorganism resistance to aminoglycoside antibiotics (by the example of gentamicin). The article represents the research results of the acylated derivatives of ellagic acid and its supramolecular derivatives with β-cyclodextrins. The structure of the compounds synthesized has been confirmed by NMR1H -spectroscopy and HPLC with a multiwave ultraviolet-detector. It has been shown that in sub-effective concentrations (0.001% solution) in the absence of direct antimicrobial and antifungal properties, tetrasuccinyl-digallic acid (5) reveals the ability to reduce sensitivity to gentamicin in polyresistant strains of microorganisms and sensitivity to nystatin in fungi resistant to it. It has been found that clathrate complexes of ellagotanin with β-cyclodextrins have no antiresistant activity, as well as other acylated derivatives of ellagic acid.