Pathogenic Gram-negative Bacteria Research Articles

Tenacibaculum dicentrarchi produce outer membrane vesicles (OMV) that are associated with the cytotoxic effect in rainbow trout head kidney macrophages.

Tenacibaculum dicentrarchi is the second most important pathogen in Chilean salmon farming. This microorganism causes severe skin lesions on the body surface of farmed fish. The bacterium can also adhere to surfaces and form biofilm, survive in fish skin mucus, and possess different systems for iron acquisition. However, the virulence mechanisms are still not fully elucidated. Outer membrane vesicles (OMV) are nanostructures released by pathogenic Gram-negative bacteria during growth, but none has been described yet for T. dicentrarchi. In this study, we provide the first reported evidence of the fish pathogen T. dicentrarchi producing and releasing OMV from 24 h after incubation, increasing thereafter until 120 h. Analyses were conducted with T. dicentrarchi TdCh05, QCR29, and the type strain CECT 7612T . The OMV sizes, determined via scanning electron microscopy, ranged from 82.25 nm to 396.88 nm as per the strain and incubation time point (i.e., 24 to 120 h). SDS-PAGE revealed that the number of protein bands evidenced a drastically downward trend among the T. dicentrarchi strains. In turn, the OMV shared five proteins (i.e., 22.2, 31.9, 47.7, 56.3, and 107.1 kDa), but no protein pattern was identical. A heterogeneous amount of protein, RNA, and DNA were obtained, depending on the time at which OMV were extracted. Purified OMV were biologically active and induced a cytotoxic effect in macrophage-enriched cell cultures from rainbow trout (Oncorhynchus mykiss) head kidneys. This is the first step towards understanding the role that OMV could play in the pathogenesis of T. dicentrarchi.

Neisseria Gonorrhoeae: Its Dominant Properties to Establish Contact and Attachment that Facilitate Epithelial Invasion and Colonization From a Biochemist Perspective

Aim: to discuss recent epidemiology of gonorrhoea and updates regarding its pathogenesis with a focus on biochemical aspects of contact and adhesion that preceded its epithelial invasion.
 Discussion: Neisseria gonorrhoeae, a Gram-negative obligate human pathogenic bacterium, infects human epithelial cells and causes sexually transmitted diseases in both males and females Gonorrhea rates are rising in many countries. It could lead to long-term health problems and even infertility. Vulnerable groups including men who have sex with men and sex workers appear to bear disproportionate burdens of gonorrhea. As N. gonorrhoeae advances through the steps of disease formation and pathogenesis (transmission, adherence, colonization and invasion, and also immune evasion), the bacterium expresses some virulence factors to facilitate its survival and replication; while at the same time keeping its existence barely invasive and almost undiscoverable by active immune cells. Adherence to epithelial cells becomes the first event that precedes invasion.
 Conclusion: Adhesion of gonococci to mucosal epithelial cells appears to be a critical step in the pathogenesis of gonococcal infection. Gonococci can adhere to a variety of human cells. Gonorrhea has multiple surface proteins that facilitate adhesion. N. gonorrhoeae utilize type IV pili and Opa, opacity-associated proteins, surface proteins involved in cellular attachment that preceded its invasion, Lipooligosaccharides (LOS) and also the outer membrane protein porin (POrB).

Computational and Experimental Investigation of Antibacterial Properties of Some Fluorinated Thioureas

Herein, fluorinated thioureas with various substituents (F1-F3) were synthesized and characterized spectroscopically and analytically to investigate their properties as antibacterial agents. Prior to experimental studies, Density Functional Theory (DFT) modeling was performed at B3LYP/6-31G (d,p) to complement and offer comparative overview with the experimental findings. With regards to all theoretical analyses, the Frontier Molecular Orbital (FMO) demonstrated a desirable low HOMO-LUMO gap for all three designated derivatives, particularly F1 (1.60 eV), which is consistent with its GCRDs values, namely its high electronegativity and low hardness values that corresponds to low LUMO energy, suggesting higher bacterial activity. Agar diffusion assay was used for the preliminary screening of in-vitro antibacterial activity against pathogenic Gram-positive and Gram-negative bacteria. All three derivates successfully inhibit at least two bacterial strains (B. cereus, S. aureus), but none were able to penetrate into E. coli. The highest inhibition rate was exhibited by F3 at 33.33 ± 0.71% against S. aureus, whilst F1 only managed to gain 29.63 ± 0.00% of inhibition rate for the same bacterium. Meanwhile, the lowest penetration rate was recorded at 1.53 ± 0.35% by F2 against B. cereus.

Generation and Investigation of FE2O3 Nanoparticles’ Optical Properties and Antibacterial Activity Using Pulsed Laser Ablation in Deionized Water

The synthesis of metallic nanoparticles has ignited intense interest over the last decade due to their unique properties, which make them applicable in a variety of fields of science and technology. In this study, iron oxide nanoparticles (Fe2O3 NPs) were synthesized, characterized, and their antibacterial activity was determined using a low-cost and simple method. Method: In this paper, magnetic nanoparticles were synthesized by a physical method of pulsed laser ablation in a liquid medium with 600 mJ energy. Magnetic Fe2O3 NPs were prepared by using a 1064 nm Nd-YAG laser and bombarding the [Formula: see text]-Fe2O3 target in a deionized aqueous solution. An absorption spectrometer was used to study the optical properties and structures of nanoparticles, the prepared nanoparticles were analyzed using Fourier transform infrared (FTIR) spectroscopy. Besides, utilizing FESEM, the morphological and surface characteristics were examined. The antibacterial activity of Fe2O3 NPs against several pathogenic Gram-positive and Gram-negative bacteria isolated from various human infection sites was investigated using a disk diffusion experiment with laser pulse lengths of (100 nm, 150 nm, 200 nm and 300 nm) with the same laser energy. Result: The results showed the success of this method as the first sign that can be inferred from the change in the color of the solution to prepare nanoparticles. Fe2O3 NPs are mostly spherical in shape, with nano sizes ranging between 66.7 nm and 9.5 nm. The energy gap was found to be in the range of 1.79–2.39 eV for almost all nanoparticles prepared. Fe2O3 NPs were formed in almost spherical surface morphology with regular size and shape. The results showed their effect on stopping the growth of bacteria of type E. coli maximum zone of inhibition of 20 mm and 19 mm of S. epidermidis, 18 mm of type S. mutans and 16 mm of P. aeruginosa. As a result, in this study, it was shown that Fe2O3 NPs produced in water by laser ablation method can be used as antibacterial agents due to these properties.

Vitex negundo and Euphorbia milii leaf extracts aided green synthesis of copper oxide nanostructures for effective inactivation of pathogenic bacteria

A facile and green route was followed to synthesize copper oxide (CuO) nanostructures using Vitex negundo and Euphorbia milii leaf extracts. Structural, elemental, chemical, spectral and morphological features of the CuO nanostructures were determined by XRD, EDS, FTIR spectroscopy, UV–Vis spectrophotometer and FESEM, respectively. The CuO nanostructures disclosed considerably higher antibacterial activity against selected Gram-positive and Gram-negative pathogenic bacteria over commercial antibiotic (Vancomycin). This enhanced antibacterial activity could be credited to the generation of more ROS associated with oxygen vacancy and size of the CuO nanostructures. Particularly, hierarchical CuO nanoparticles/nanorods prepared using V. negundo leaf extract exhibited the highest antibacterial activity due to their lower size range.

Antibacterial and antibiofilm activity of nanocomposite (BJ-Ag0.6@GO0.4) synthesized from animal-derived liquid waste against ESKAPE pathogen

Herein, Ag nanoparticles were prepared by fish bile juice as a reducing and capping agent. Then Ag/ graphene oxide (GO) nanocomposite was fabricated. The nanocomposite (BJ-Ag0.6 @GO0.4) showed remarkable antibacterial activities against Gram-positive and Gram-negative pathogenic bacteria. The MIC values of the nanocomposite were recorded as 10 and 8 µg/mL for S.aureus (MTCC 1430) and P. auregenosa (MTCC-1688), respectively. Antibacterial mechanism study of the synthesized nanocomposite excavates that, BJ-Ag0.6 @GO0.4 caused extensive membrane damage and cytoplasmic leakage, leading to cell’s metabolic arrest. Increased ROS generation is observed, causing disruption in the antioxidant defense mechanism of the bacterial cell. The synthesized nanocomposite also showed excellent destruction of preformed biofilm up to 96.52% and 61.23% for S.aureus (MTCC 1430) and P. auregenosa (MTCC-1688), respectively, at MIC concentration of the nanocomposite.

The Importance of Visceral Hypersensitivity in Irritable Bowel Syndrome-Plant Metabolites in IBS Treatment.

The visceral stimuli from the digestive tract are transmitted via afferent nerves through the spinal cord to the brain, where they are felt as pain. The overreaction observed in the brain of irritable bowel syndrome (IBS) patients may be due to increased peripheral sensitivity to stimuli from the gastrointestinal tract. Although the exact pathway is uncertain, attenuation of visceral hypersensitivity is still of interest in treating IBS. It has been shown that stress stimulates the sympathetic nervous system while inhibiting the vagus nerve (VN). In addition, stress factors lead to dysbiosis and chronic low-grade inflammation of the intestinal mucosa, which can lead to lower gastrointestinal visceral hypersensitivity. Therefore, an important goal in the treatment of IBS is the normalization of the intestinal microflora. An interesting option seems to be nutraceuticals, including Terminalia chebula, which has antibacterial and antimicrobial activity against various pathogenic Gram-positive and Gram-negative bacteria. Additionally, short-term transcutaneous vagus nerve stimulation can reduce the stress-induced increase in intestinal permeability, thereby reducing inflammation. The conducted studies also indicate a relationship between the stimulation of the vagus nerve (VN) and the activation of neuromodulatory networks in the central nervous system. Therefore, it seems reasonable to conclude that a two-way action through stimulating the VN and using nutraceuticals may become an effective therapy in treating IBS.

Effect of steam sterilisation on physico-chemical properties of antibacterial covalently cross-linked ε-polylysine/hyaluronic acid hydrogels

Sterilisation of implantable biomaterials such as hydrogels remains a key step towards their clinical translation. Standard sterilisation methods can significantly alter hydrogels' physicochemical and biological performance. Previously, we developed composite hydrogels based on ε-polylysine (ε-PL) and hyaluronic acid (HA). The developed hydrogels demonstrated promising antibacterial activity and in vitro cell viability and their variable properties depending on the chosen ε-PL to HA ratio. In this study, we fabricated a series of chemically cross-linked ԑ-PL/HA hydrogels with expanded ԑ-PL to HA mass ratios.Using small-angle X-ray scattering (SAXS), we unravelled the topological differences between physically and chemically crosslinked hydrogels. We then selected the chemically crosslinked hydrogel ԑ-PL/HA series of 60:40 wt%, 70:30 wt%, and 80:20 wt% ratios, with similar network topologies, to evaluate the impact of steam sterilisation on their physicochemical and viscoelastic properties. The antibacterial activity of the sterilized hydrogels was also evaluated against Gram-negative and Gram-positive bacteria. Our results showed that steam sterilisation minimally affects structure and physicochemical properties of ԑ-PL/HA hydrogels. Furthermore, the developed hydrogel ԑ-PL/HA series of 60:40 wt%, 70:30 wt%, and 80:20 wt% ratios showed pronounced antibacterial activity against Gram-negative and Gram-positive pathogenic bacteria. We expect our results will contribute to the growing understanding of using sterilisation methods for antibacterial hydrogels that have the potential for wider tissue engineering applications.

Controlling bacterial biofilm formation by native and methylated lupine 11S globulins.

The antibacterial and anti-biofilm activities of the 11S globulins isolated from lupin seeds (Lupinus termis), and its methylated derivative (M11S), were investigated against seven pathogenic gram-positive and gram-negative bacteria. The MIC of 11S ranged from 0.1 to 4.0 μg/ml against 0.025 to 0.50 μg/ml for M11S, excelling some specific antibiotics. The MICs of M11S were 40-80 times lower than some specific antibiotics against gram-positive bacteria and 2-60 times lower than some specific antibiotics against gram-negative bacteria. One MIC of 11S and M11S highly reduced the liquid growth of all tested bacteria during 24 h at 37°C. They also inhibited biofilm formation by 80%-86% and 85%-94%, respectively (gram-positive), and 29%-44% and 43%-50%, respectively (gram-negative). M11S prevented biofilm formation by gram-positive bacteria at minimum biofilm inhibitory concentration (MBIC), 0.025-0.1 μg/ml against 0.1-0.5 μg/ml for gram-negative bacteria, i.e., 4-20 times and 4-7 times anti-biofilm inhibitory action compared with 11S, respectively. Biofilm formation of two bacteria revealed no adhered cells on glass slides for 24 h at 37°C, i.e., was entirely prevented by one MBIC of 11S and M11S. Scanning electron microscopy indicated microbial biofilm deformation under the action of 11S and M11S, indicating their broad specificity and cell membrane-targeted action.

Whole genome sequence analysis of Aeromonas-infecting bacteriophage AHPMCC7, a new species of genus Ahphunavirus and its application in Litopenaeus vannamei culture

Aeromonas hydrophila, a Gram-negative pathogenic bacterium, is responsible for huge economic losses in aquaculture. In this study, we evaluated the efficacy of bacteriophage AHPMCC7 which was isolated by using A. hydrophila MTCC 1739 as a host. This bacteriophage exhibited 10 min latent period and burst size was 275. In liquid culture, bacteriophage AHPMCC7 could completely lyse A. hydrophila MTCC 1739 after 2 h. AHPMCC7 genome was 42,277 bp long with 58.9% G + C content. The genome consisted of 48 CDSs and no tRNA. The comparative genomic analyses clearly implied that AHPMCC7 might represent a novel species of the genus Aphunavirus under Autographiviridae family. Bacteriophage AHPMCC7 could survive at broad pH (3–10), temperature (4–37 °C), and salinity (0–40 ppt). In aquarium trial, AHPMCC7 could control A. hydrophila MTCC 1739 without affecting the survivability of Litopenaeus vannamei. Clearly, the bacteriophage AHPMCC7 might be used in shrimp aquaculture as a biocontrol agent.

Green synthesis of Multifunctional Zinc oxide Nanoparticles from Cordia myxa gum; and their Catalytic Reduction of Nitrophenol, Anticancer and Antimicrobial Activity

In situ exfoliated natural polysaccharide Cordia myxa (CMX) is used to promote the utilization of zinc-oxide nanoparticles for eco-friendly catalytic hydrogenation of p-nitrophenol (p-NP) and microbial growth inhibition. Polysaccharide-mediated biosynthetic nanocomposite materials are interesting because they are cheap, green, and environmentally friendly. This study uses CMX gum as a bioreduction to produce multifunctional, environmentally friendly zinc-oxide nanocomposites (ZnO NPs). The process involves a low reaction time and temperature and utilizes CMX as a reducing and stabilizing agent. The structural, morphological, and optical properties of the CMX-ZnO nanocomposite were characterized. The biosynthetic CMX-ZnO NPs exhibited robust catalytic activity and recycling capacity for rapidly oxidizing hazardous p-NPs. The complete reduction of 4-NP to CMX-ZnO NPs in excess NaBH4 was achieved within 15 min, with recyclability and pseudo-first-order kinetics with a rate constant of 0.2571 min−1. Additionally, human colon cancer (HCT116) and 3T3L1 cell lines were remarkably sensitive to the cytotoxic effects of ZnO nanoparticles. CMX-ZnO NPs exhibited potent antibacterial properties against human pathogenic gram-positive and gram-negative bacteria (Bacillus, Salmonella, E. coli, and Pseudomonas aeruginosa) based on the zone of inhibition measured by the disc-diffusion method. The significant antibacterial activity of CMX-ZnO NPs can overcome the current limitations associated with removing water-soluble organic pollutants and microbiological contaminants for long-term environmental sustainability.

Biosynthetic gene cluster signature profiles of pathogenic Gram-negative bacteria isolated from Egyptian clinical settings.

Biosynthetic gene clusters (BGCs) are a subset of consecutive genes present within a variety of organisms to produce specialized metabolites (SMs). These SMs are becoming a cornerstone to produce multiple medications including antibacterial and anticancer agents. Natural products (NPs) also play a pivotal role in enhancing the virulence of ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.), which represent a global health threat. We aimed to sequence and computationally analyze the BGCs present in 66 strains pertaining to three different ESKAPE pathogenic species: 21 A. baumannii, 28 K. pneumoniae, and 17 P. aeruginosa strains recovered from clinical settings in Egypt. DNA was extracted using QIAamp DNA Mini kit and Illumina NextSeq 550 was used for whole-genome sequencing. The sequences were quality-filtered by fastp and assembled by Unicycler. BGCs were detected by antiSMASH, BAGEL, GECCO, and PRISM, and aligned using Clinker. The highest abundance of BGCs was detected in P. aeruginosa (590), then K. pneumoniae (146) and the least in A. baumannii strains (133). P. aeruginosa isolates shared mostly the non-ribosomal peptide synthase (NRPS) type, K. pneumoniae isolates shared the ribosomally synthesized and post-translationally modified peptide-like (RiPP-like) type, while A. baumannii isolates shared the siderophore type. Most of the isolates harbored non-ribosomal peptide (NRP) BGCs with few K. pneumoniae isolates encoding polyketide BGCs. Sactipeptides and bottromycin BGCs were the most frequently detected RiPP clusters. We hypothesize that each species' BGC signature confers its virulence. Future experiments will link the detected clusters with their species and determine whether the encoded SMs are produced and cause their virulence. IMPORTANCE Our study analyzes the biosynthetic gene clusters (BGCs) present in 66 assemblies from clinical ESKAPE pathogen isolates pertaining to Acinetobacter baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa strains. We report their sequencing and assembly followed by the analysis of their BGCs using several bioinformatics tools. We then focused on the most abundant BGC type in each species and we discussed their potential roles in the virulence of each species. This study is pivotal to further build on its experimental work that deciphers the role in virulence, possible antibacterial effects, and characterization of the encoded specialized metabolites (SMs). The study highlights the importance of studying the "harmful" BGCs and understanding the pathogenicity and virulence of those species, as well as possible benefits if the SMs were used as antibacterial agents. This could be the first study of its kind from Egypt and would shed light on BGCs from ESKAPE pathogens from Egypt.

Development of an antibody fused with an antimicrobial peptide targeting Pseudomonas aeruginosa: A new approach to prevent and treat bacterial infections.

The increase in emerging drug resistant Gram-negative bacterial infections is a global concern. In addition, there is growing recognition that compromising the microbiota through the use of broad-spectrum antibiotics can impact long term patient outcomes. Therefore, there is the need to develop new bactericidal strategies to combat Gram-negative infections that would address these specific issues. In this study, we report and characterize one such approach, an antibody-drug conjugate (ADC) that combines (i) targeting the surface of a specific pathogenic organism through a monoclonal antibody with (ii) the high killing activity of an antimicrobial peptide. We focused on a major pathogenic Gram-negative bacterium associated with antibacterial resistance: Pseudomonas aeruginosa. To target this organism, we designed an ADC by fusing an antimicrobial peptide to the C-terminal end of the VH and/or VL-chain of a monoclonal antibody, VSX, that targets the core of P. aeruginosa lipopolysaccharide. This ADC demonstrates appropriately minimal levels of toxicity against mammalian cells, rapidly kills P. aeruginosa strains, and protects mice from P. aeruginosa lung infection when administered therapeutically. Furthermore, we found that the ADC was synergistic with several classes of antibiotics. This approach described in this study might result in a broadly useful strategy for targeting specific pathogenic microorganisms without further augmenting antibiotic resistance.

Structural analysis of a bacterial UDP-sugar 2-epimerase reveals the active site architecture before and after catalysis

The sugar, 2,3-diacetamido-2,3-dideoxy-d-mannuronic acid, was first identified ∼40 years ago in the O-antigen of Pseudomonas aeruginosa O:3,a,d. Since then, it has been observed on the O-antigens of various pathogenic Gram-negative bacteria including Bordetella pertussis, Escherichia albertii, and Pseudomonas mediterranea. Previous studies have established that five enzymes are required for its biosynthesis beginning with UDP-N-acetyl-d-glucosamine (UDP-GlcNAc). The final step in the pathway is catalyzed by a 2-epimerase, which utilizes UDP-2,3-diacetamido-2,3-dideoxy-d-glucuronic acid as its substrate. Curious as to whether this biochemical pathway is found in extreme thermophiles, we examined the published genome sequence for Thermus thermophilus HB27 and identified five open- reading frames that could possibly encode for the required enzymes. The focus of this investigation is on the open reading frame WP_011172736, which we demonstrate encodes for a 2-epimerase. For this investigation ten high resolution X-ray crystallographic structures were determined to resolutions of 2.3 Å or higher. The models have revealed the manner in which the 2-epimerase anchors its UDP-sugar substrate as well as its UDP-sugar product into the active site. In addition, this study reveals for the first time the manner in which any sugar 2-epimerase can simultaneously bind UDP-sugars in both the active site and the allosteric binding region. We have also demonstrated that the T. thermophilus enzyme is allosterically regulated by UDP-GlcNAc. Whereas the sugar 2-epimerases that function on UDP-GlcNAc have been the focus of past biochemical and structural analyses, this is the first detailed investigation of a 2-epimerase that specifically utilizes UDP-2,3-diacetamido-2,3-dideoxy-d-glucuronic acid as its substrate.

Biologically active selenium nanoparticles composited with Bacillus licheniformis extracellular polymeric substances fermented from cane molasses

Organic selenium nanoparticles (SeNPs) are attracting great interest due to their unique properties and potential applications in the food industry. In this study, a novel form of SeNPs was synthesized by using Bacillus licheniformis CGMCC 2876 extracellular polymeric substances (EPS) fermented from waste cane molasses. The morphology of EPS-SeNPs was spherical with an average size of 141.8 nm and a zeta potential of −51.77 mV, and the concentration of Se reached 107.43 mg/g. Additionally, the EPS-SeNPs exhibited good stability in aqueous solution for 7 days and were resistant to the digestion of simulated gastric fluid. The zero-valent Se composition and C–O⋯Se asymmetric stretch were characterized in the EPS-SeNPs. A synthesis mechanism of EPS-SeNPs was hypothesized. Moreover, the biological activities of the EPS-SeNPs synthesized from EPS were evaluated. The EPS-SeNPs exhibit antioxidant activity in scavenging ·OH, DPPH and ·O2− free radicals with abilities <90% in the range of 0.25–4 g/L and inhibit 50% of formed radicals (IC50) values of 1.18, 13.71 and 1.93 g/L, respectively. EPS-SeNPs also exhibit excellent antibacterial activity, which could inhibit the growth of intestinal pathogenic gram-positive and gram-negative bacteria (>80%) in vitro with the EPS-SeNPs concentrations higher than 1000 μg/mL.

Phosphoethanolamine Transferases as Drug Discovery Targets for Therapeutic Treatment of Multi-Drug Resistant Pathogenic Gram-Negative Bacteria.

Antibiotic resistance caused by multidrug-resistant (MDR) bacteria is a major challenge to global public health. Polymyxins are increasingly being used as last-in-line antibiotics to treat MDR Gram-negative bacterial infections, but resistance development renders them ineffective for empirical therapy. The main mechanism that bacteria use to defend against polymyxins is to modify the lipid A headgroups of the outer membrane by adding phosphoethanolamine (PEA) moieties. In addition to lipid A modifying PEA transferases, Gram-negative bacteria possess PEA transferases that decorate proteins and glycans. This review provides a comprehensive overview of the function, structure, and mechanism of action of PEA transferases identified in pathogenic Gram-negative bacteria. It also summarizes the current drug development progress targeting this enzyme family, which could reverse antibiotic resistance to polymyxins to restore their utility in empiric therapy.

Structures of the Insecticidal Toxin Complex Subunit XptA2 Highlight Roles for Flexible Domains.

The Toxin Complex (Tc) superfamily consists of toxin translocases that contribute to the targeting, delivery, and cytotoxicity of certain pathogenic Gram-negative bacteria. Membrane receptor targeting is driven by the A-subunit (TcA), which comprises IgG-like receptor binding domains (RBDs) at the surface. To better understand XptA2, an insect specific TcA secreted by the symbiont X. nematophilus from the intestine of entomopathogenic nematodes, we determined structures by X-ray crystallography and cryo-EM. Contrary to a previous report, XptA2 is pentameric. RBD-B exhibits an indentation from crystal packing that indicates loose association with the shell and a hotspot for possible receptor binding or a trigger for conformational dynamics. A two-fragment XptA2 lacking an intact linker achieved the folded pre-pore state like wild type (wt), revealing no requirement of the linker for protein folding. The linker is disordered in all structures, and we propose it plays a role in dynamics downstream of the initial pre-pore state.

Antibacterial Properties of Methanolic Leaf Extracts of Melia azedarach L. against Gram-Positive and Gram-Negative Pathogenic Bacteria.

Melia azedarach L., a Meliaceae family tree, is widely used in traditional folkloric medicine for its pharmaceutical properties. In the present study, we investigated the phytochemical composition of four methanolic leaf extracts of M. azedarach of various origins (Algeria and Tunisia) using high-performance liquid chromatography (HPLC). The antibacterial efficacy and mechanisms of action against Gram-positive and Gram-negative pathogenic microorganisms were then evaluated. Our findings revealed a presence of phenolic acids and flavonoids, such as gallic acid, chlorogenic acid, caffeic acid, hyperoside, isoquercetin, quercetin, and isorhamnetin both in Algerian and Tunisian localities, with an abundance of phenolic acids compared to flavonoids. Additionally, the studied extracts exhibit a broad spectrum of antibacterial activities, with MIC values ranging from 31.25 mg/mL to 125 mg/mL. Methanolic leaf extracts of M. azedarach from Algeria exhibited more potent biofilm eradication, with a percentage of inhibition reaching 72.17% against the S. aureus strain. Furthermore, inhibitory concentrations of tested substances, particularly the extract from the Relizane area, were capable of disrupting the membrane integrity of the treated bacteria as well as producing oxidative stress through ROS generation. Likewise, our results reveal that plant extract induces lipid peroxidation by raising MDA levels in comparison to untreated cells, particularly with the plant extract of Blida. M. azedarach extracts also reduced the synthesis of antioxidant enzymes (CAT and SOD). Our findings illustrate that M. azedarach remains a plant with significant antibacterial potential and distinct mechanisms of action that are closely related to the origins of this specimen.

Silver nanoparticles with plasma-polymerized hexamethyldisiloxane coating on 3D printed substrates are non-cytotoxic and effective against respiratory pathogens.

Due to the emerging resistance of microorganisms and viruses to conventional treatments, the importance of self-disinfecting materials is highly increasing. Such materials could be silver or its nanoparticles (AgNPs), both of which have been studied for their antimicrobial effect. In this study, we compared the biological effects of AgNP coatings with and without a plasma-polymerized hexamethyldisiloxane (ppHMDSO) protective film to smooth silver or copper coatings under three ambient conditions that mimic their potential medical use (dry or wet environments and an environment simulating the human body). The coatings were deposited on 3D printed polylactic acid substrates by DC magnetron sputtering, and their surface morphology was visualized using scanning electron microscopy. Cytotoxicity of the samples was evaluated using human lung epithelial cells A549. Furthermore, antibacterial activity was determined against the Gram-negative pathogenic bacterium Pseudomonas aeruginosa PAO1 and antiviral activity was assessed using human rhinovirus species A/type 2. The obtained results showed that overcoating of AgNPs with ppHMDSO creates the material with antibacterial and antiviral activity and at the same time without a cytotoxic effect for the surrounding tissue cells. These findings suggest that the production of 3D printed substrates coated with a layer of AgNPs-ppHMDSO could have potential applications in the medical field as functional materials.

Conformational investigation of the asymmetric periplasmic domains of E. coli LptB2FGC using SDSL CW EPR spectroscopy.

The majority of pathogenic Gram-negative bacteria benefit from intrinsic antibiotic resistance, attributed primarily to the lipopolysaccharide (LPS) coating of the bacterial envelope. To effectively coat the bacterial cell envelope, LPS is transported from the inner membrane by the LPS transport (Lpt) system, which comprises seven distinct Lpt proteins, LptA-G, that form a stable protein bridge spanning the periplasm to connect the inner and outer membranes. The driving force of this process, LptB2FG, is an asymmetric ATP binding cassette (ABC) transporter with a novel architecture and function that ejects LPS from the inner membrane and facilitates transfer to the periplasmic bridge. Here, we utilize site-directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy to probe conformational differences between the periplasmic domains of LptF and LptG. We show that LptC solely interacts with the edge β-strand of LptF and does not directly interact with LptG. We also quantify the interaction of periplasmic LptC with LptF. Additionally, we show that LPS cannot enter the protein complex externally, supporting the unidirectional LPS transport model. Furthermore, we present our findings that the presence of LPS within the LptB2FGC binding cavity and the membrane reconstitution environment affect the structural orientation of the periplasmic domains of LptF and LptG, but overall are relatively fixed with respect to one another. This study will provide insight into the structural asymmetry associated with the newly defined type VI ABC transporter class.