Protein Leakage Research Articles

Bioactivities of secondary metabolites of two actinomycetes Streptomyces parvulus GloL3, and Streptomyces lienomycini SK5, endophytes of two Indian medicinal herbs- Globba marantina L. and Selaginella kraussiana (Kunze) A. Braun.

Endophytic actinomycetes are potential sources of novel pharmaceutically active metabolites, significantly advancing natural product research. In the present investigation, secondary metabolites from two endophytic actinomycetes, Streptomyces parvulus GloL3, and Streptomyces lienomycini SK5, isolated from medicinal plant taxa, Globba marantina, and Selaginella kraussiana, exhibited broad-spectrum bioactivity. Ethyl Acetate (EA) extract of SK5 showed antimicrobial activity against nine human pathogens, including Methicillin-resistant Staphylococcus aureus (MRSA), Candida tropicalis, and C. albicans, with a minimum microbicidal concentration (MMC) of 50-300µg mL-1. It healed the MRSA-mediated wounds in Swiss albino mice in vivo. EA extracts dissociate the pathogenic cell membranes and cause leakage of biomacromolecules-nucleic acid, protein, and potassium ions. Also, critical housekeeping enzymes involved in the cellular respiratory mechanisms of the pathogens were blocked. GloL3 has antioxidative potentialities against DPPH, ABTS, FRAP, and H2O2 free radical generators with an IC50 value of 21.18 ± 0.33, 43.58 ± 0.91, 88.24 ± 1.24, and 111.03 ± 6.42µg mL-1. It improves the enzymatic antioxidant parameters in treated peritoneal macrophage cells of Swiss albino mice. Constituents of the EA extracts of GloL3 and SK5 are bactobolin, actinobolin, 5-(2-aminoethyl)-1H imidazole-2-carbaldehyde, isovaleric acid, fulvic acid, phenol, 4-[2-(methylamino) ethyl]-, eicosanoic acid, heptadecanoic acid, etc. The present findings suggest that metabolites from the endophytes of medicinal plants hold potent pharmaceutical utilities.

Evaluation of extracts from Phyllostachys makinoi for their antibacterial and accelerated wound healing potential

Phyllostachys makinoi, an endemic bamboo species in Taiwan, is underutilized, despite its rich forest resources. Known for its antioxidant, anti-inflammatory, and antibacterial properties, this study explores the antimicrobial, anti-inflammatory, and wound-healing activities of P. makinoi extracts. The antibacterial potential of P. makinoi extracts was first evaluated using the agar diffusion method, along with minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assays. Subsequently, electron microscopy and a conductivity meter were employed to assess whether P. makinoi extracts exert antibacterial effects by disrupting bacterial cell structures. Finally, the anti-inflammatory and cell proliferation-promoting effects of P. makinoi extracts were assessed in RAW264.7 and CCD-1112Sk cell models. The MIC and MBC of the P. makinoi water extracts against two multidrug-resistant Staphylococcus aureus strains were 4 and 16–32 mg/mL, respectively, while those for ethanol extracts were 2 and 32 mg/mL, respectively. In the time-kill assay, both strains were killed after treatment with extracts for 12 and 18 h. Scanning electron microscopy revealed that the bacterial cells treated with the extracts appeared rough, ruptured, and shriveled. The extracts disrupted the cell membranes, causing electrolyte, protein, and nucleic acid leakage, leading to cell death. Additionally, P. makinoi extracts reduced lipopolysaccharide-induced nitric oxide, tumor necrosis factor-α, interleukin (IL)-1β, and IL-6 levels in RAW264.7 cells, and promoted wound healing by enhancing CCD-1112Sk cell proliferation. In conclusion, P. makinoi water and ethanol extracts demonstrated antibacterial and anti-inflammatory effects, showing potential for treating wound infections and accelerating healing, making them promising candidates for wound-healing therapies.

Inactivation activity and mechanism of high-voltage pulsed electric fields combined with antibacterial agents against Alicyclobacillus spp. in apple juice.

Alicyclobacillus spp. are crucial factors affecting the quality of fruit juice, so it is very important to control their contamination. In this study, the inactivation activity and mechanism of high-voltage pulsed electric fields (HVPEF) combined with antibacterial agents against Alicyclobacillus spp. in apple juice were investigated. It was found that under the optimal conditions of electric field strength 9.6kV/cm, treatment time 20min, frequency 1000Hz, and duty ratio 50%, HVPEF treatment could reduce bacteria by 1.89-4.76 log CFU/mL. Moreover, the inactivation activities of six antibacterial agents (propyl paraben, glycerol monocaprate, octyl gallate, heptyl paraben, nisin, carvacrol) alone and their combination with HVPEF were further investigated. The results showed that with the combined treatment, the minimum bactericidal concentrations of carvacrol, nisin, and heptyl paraben were reduced by >50% to 1mg/mL, 10IU/mL, and 0.02mg/mL, respectively. Based on this, the most resistant strain of A. acidoterrestris (DSM 3922) was identified to elucidate the inactivation mechanism. It was demonstrated that the antibacterial process could alter the permeability and fatty acid composition of the cell membrane, causing the cells to deform and shrink, leading to leakage of intracellular proteins, and also affect the synthesis of ROS and ATP, ultimately resulting in bacterial death. In addition, the various treatments had no significant effect on the soluble solids content, titratable acid, soluble sugar content, organic acids and aroma components of apple juice. The combination of HVPEF treatment and antibacterial agents could effectively maintain the quality of apple juice.

Understanding antimicrobial activity of biogenic selenium nanoparticles and selenium/chitosan nano-incorporates via studying their inhibition activity against key metabolic enzymes.

Microbial metabolic enzymes play a crucial role in several biological processes that have a significant impact on growth and proliferation. Therefore, inhibiting specific key metabolic enzymes can be an applicable approach for developing antimicrobial agents that selectively target pathogens. In the current study, selenium nanoparticles (Se NPs) extracellularly biosynthesized by Nocardiopsis sp. MAR13 and Se NPs incorporated with nano-chitosan (NCh) (Se/Ch-nano-incorporate) were evaluated for their antimicrobial activity against various microbial pathogens such as Salmonella typhi, Proteus vulgaris, Staphylococcus aureus, Escherichia coli, Aspergillus flavus, Aspergillus niger, Rhizoctonia sp., Candida albicans ATCC10231. The synthesized Se NPs, and Se/Ch-nano-incorporate were characterized by UV-Vis., FTIR, HRTEM, SEM, EDX, DLS, Zeta potential, and XRD. Additionally, their inhibition activity against microbial metabolic key enzymes, including phosphoglucose isomerase (PGI), pyruvate dehydrogenase (PDH), glucose-6-phosphate dehydrogenase (G6PDH), and nitrate reductase (NR), was assessed. The impact on protein leakage from bacterial cell membranes was also evaluated as a potential mechanism of antimicrobial action. On the other hand, MCF-7 and A549 tumor cell lines, as well as WI-38 normal cell lines, were used to assess their cytotoxic activity. It was found that Se NPs were spherical with a diameter range of 60.2 to 120.2nm. In contrast, Se/Ch-nano-incorporate had a roughly spherical shape with a diameter range of 21.4 to 32.7nm and substantially higher stability. Both synthesized agents exhibited strong antimicrobial activity against the most tested microbial pathogens with substantial inhibitory effect on the tested enzymes and notable protein leakage. Furthermore, they showed potent anticancer activity against both tumor cell lines with low cytotoxicity against WI-38 normal cell line. Consequently, Se NPs and Se/Ch-nano-incorporate are highly recommended to be employed as antimicrobial and anticancer agents with promised biosafety, eco-friendliness, and efficacy.

‘Breaking the biofilm barrier: vitamin C as a novel strategy against multidrug-resistant salmonella Typhimurium’

Abstract The emergence of multidrug-resistant (MDR), biofilm-forming strains of Salmonella Typhimurium poses a significant threat to food safety due to their persistence on food surfaces and equipment, complicating eradication efforts and increasing the risk of contamination. This study explored the antibiofilm activity of Vitamin C (VC) against MDR biofilm-forming Salmonella Typhimurium. Vitamin C inhibited S. Typhimurium biofilms in a dose-dependent manner, with a minimum inhibitory concentration of 125 mM. Treatment with Vitamin C significantly disrupted the bacterial biofilm structure, creating cannonball-like pores in the cell wall as observed through electron microscopy. The antibiofilm mechanism of Vitamin C was mediated by reactive oxygen species (ROS) generation, leading to sugar and protein leakage, reduced exopolysaccharide (EPS) production, and decreased quorum sensing (QS) activity. Transcriptional analysis revealed that Vitamin C downregulated expression of biofilm-related genes responsible for initial cellular attachment and EPS production (fimA, adrA) and upregulated genes associated with adhesion and multidrug efflux pumps (csgA, ipfE, and arcA). Importantly, the application of Vitamin C on coated Paneer significantly extended its shelf life. These findings highlight the potential of Vitamin C in combating Salmonella biofilms, enhancing food safety, and providing a framework for the development of antibiotic-independent therapies to address MDR strains of Salmonella.

Antifungal lipopeptides from the marine Bacillus amyloliquefaciens HY2-1: A potential biocontrol agent exhibiting in vitro and in vivo antagonistic activities against Penicillium digitatum.

This study aimed to clarify the antifungal activity and action mechanism of the lipopeptides from a marine Bacillus amyloliquefaciens HY2-1 against Penicillium digitatum both in vitro and in vivo. Results showed that HY2-1 lipopeptides exerted obvious inhibitions on spore germination and mycelium growth of P. digitatum. Furthermore, HY2-1 lipopeptides caused the aggravated lipid peroxidation, the decreased ergosterol biosynthesis, and the increased nucleic acid and protein leakage in P. digitatum, suggesting the damaged membrane integrity and permeability of P. digitatum. In addition, HY2-1 lipopeptides induced ROS burst and reduced the activities of antioxidant superoxide dismutase and peroxidase in P. digitatum, indicating that the intracellular redox homeostasis of P. digitatum was disturbed. In vivo biocontrol experiments showed that HY2-1 lipopeptides could effectively reduce the occurrence of green mold disease in citrus fruits artificially inoculated with P. digitatum spores. Most significantly, 4.8mg/mL of HY2-1 lipopeptides achieved 100% of biocontrol efficacy after 30days of storage and increased significantly contents of total phenols and flavonoids in citrus peels, demonstrating that HY2-1 lipopeptides inhibited green mold disease by exerting their antifungal activities and triggering fruit defense responses. This study deepens the understanding of marine Bacillus lipopeptides in the biological control of postharvest diseases.

Synergistic Effects and Mechanisms of Action of Rutin with Conventional Antibiotics Against Escherichia coli.

Rutin is a widely known plant secondary metabolite that exhibits multiple physiological functions. The present study focused on screening for synergistic antibacterial combinations containing rutin, and further explored the mechanisms behind this synergy. In vitro antibacterial test results of rutin showed that the ranges of minimum inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC) are 0.125-1 and 0.125-2 mg/mL, respectively. However, rutin and amikacin have a significant synergistic effect, with a fractional inhibitory concentration index (FICI) range of 0.1875-0.5. The time bactericidal curve proved that the combination of rutin and amikacin inhibited bacterial growth within 8 h. Scanning electron microscopy (SEM) revealed that a low-dose combination treatment could disrupt the cell membrane of Escherichia coli (E. coli). A comprehensive analysis using alkaline phosphatase (AKP), K+, and a protein leakage assay revealed that co-treatment destroyed the cell membrane of E. coli, resulting in the significant leakage of AKP, intracellular K+, and proteins. Moreover, confocal laser scanning microscopy (CLSM) and red-green cell ratio analysis indicated severe damage to the E. coli cell membrane following the co-treatment of rutin and amikacin. This study indicates the remarkable potential of strategically selecting antibacterial agents with maximum synergistic effect, which could significantly control antibiotic resistance.

Lung immune incompetency after mild peritoneal sepsis and its partial restoration by type 1 interferon: a mouse model study

BackgroundSepsis is commonly associated with acute respiratory distress syndrome (ARDS). Although the exaggerated inflammation may damage intact lung tissues, a percentage of patients with ARDS are reportedly immunocompromised, with worse outcomes. Herein, using a murine sepsis model, time-course immune reprogramming after sepsis was evaluated to explore whether the host is immunocompromised. Leukocyte kinetics in the lung tissue were evaluated in a male C57/BL6 mouse model of mild peritoneal sepsis induced by cecal ligation and puncture, with the survival rate exceeds 90%. Lung immune reactivity was evaluated by intratracheal instillation of lipopolysaccharide (LPS; 30 µg). Furthermore, the effect of interferon (IFN)-β in vivo and ex vivo was evaluated.ResultsFour days after sepsis, the lung water content remained high, even among mice in clinical recovery. While monocytes and neutrophils gradually accumulated in the lung interstitium, the inflammatory cytokine/chemokine expression levels in the lungs continued to decline. Intratracheal LPS instillation induced more leukocyte trafficking and protein leakage into the alveoli in the septic lung, indicating more severe lung injury. However, LPS stimulation-associated mRNA expression of tnf, il6, ccl2, and cxcl1 was suppressed. Intra-alveolar expression of tumor necrosis factor (TNF)-α, interleukin (IL)-6, monocyte chemoattractant protein (MCP)-1, and keratinocyte-derived cytokine (KC) was also suppressed. Monocytes isolated from the lung tissue showed an impaired response in il6, ccl2, and cxcl1 to LPS. Systemic IFN-β restored the above impaired regulator function of monocytes, as did coculturing these cells from lung tissue with IFN-β.ConclusionsHistologically accelerated inflammation and paradoxically suppressed immunological regulator signaling were observed in the early recovery phase of sepsis. This observation may provide a model for the immunologically irresponsive state that occurs in some patients with sepsis. Systemic IFN-β partly restored the post-septic immunocompromised state, indicating its therapeutic potential for the immunosuppressive state seen in some patients with sepsis/ARDS.

Constituents, Antioxidant and Antibacterial Abilities of Essential Oil of Dolichos lablab Flowers and Its Application as Food Preservative

ABSTRACTDolichos lablab flowers, as the dried buds of D. lablab L, was used as an ingredient in various dishes in China, and there were no studies on the constituents, antioxidant and antibacterial activity with the essential oil. The constituents, antioxidant and antibacterial ability of the essential oil of D. lablab flowers (EOF) obtained by hydro‐distillation (HD) method were evaluated, and its efficacy in fresh pork preservation was studied. The 36 components were recognised, accounting for 96.46% of EOF. The proportion of oxygen‐containing compounds in EOF was as high as 89.72%, of which fatty acid accounted for 65.82%. Among fatty acids, palmitic acid (33.86%), myristic acid (8.42%), lauric acid (5.42%) and 2E‐dodecenoic acid (4.46%) were identified as the major compounds. EOF had an excellent antioxidant ability. The semi‐radical scavenging concentrations (IC50) of EOF for 2,2‐Diphenyl‐1‐picrylhydrazyl (DPPH−), 2,2′‐Azinobis‐(3‐ethylbenzthiazoline‐6‐sulphonate) (ABTS+) and liposomal peroxidation were 3.09, 2.46 and 0.82 mg/mL. EOF also showed good antibacterial ability, including Bacillus subtilis, Listeria monocytogenes, Escherichia coli and Staphylococcus aureus. And the L. monocytogenes showed the highest sensitivity to EOF with the minimum inhibitory concentration (MIC) of 1.25 μg/mL and the minimum bactericidal concentration (MBC) of 2.5 μg/mL. Further research showed that EOF achieved its antibacterial effect by destroying the functional and structural integrity of cell membranes and cell walls of L. monocytogenes, leading to the leakage of alkaline phosphatase (AKP), nucleic acids and proteins, ultimately leading to L. monocytogenes death. The pork fresh‐keeping results showed that 1.0% EOF extend the shelf‐life of pork up to day 6, which was 2.00 times than the control during 7 days of refrigeration at 4°C. This study will provide the fundamental data support for future potential applications of EOF in food preservation.

Restoring Multidrug-Resistant Escherichia coli Sensitivity to Ampicillin in Combination with (-)-Epigallocatechin Gallate.

Multidrug-resistant (MDR) bacteria, especially Escherichia coli, are a major contributor to healthcare-associated infections globally, posing significant treatment challenges. This study explores the efficacy of (-)-epigallocatechin gallate (EGCG), a natural constituent of green tea, in combination with ampicillin (AMP) to restore the effectiveness of AMP against 40 isolated MDR E. coli strains. Antimicrobial activity assays were conducted to determine the minimum inhibitory concentrations (MIC) of EGCG using the standard microdilution technique. Checkerboard assays were employed to assess the potential synergistic effects of EGCG combined with AMP. The pharmacodynamic effects of the combination were evaluated through time-kill assays. Outer membrane disruption was analyzed by measuring DNA and protein leakage and with assessments using N-phenyl-1-naphthylamine (NPN) and rhodamine 123 (Rh123) fluorescence dyes. Biofilm eradication studies involved biofilm formation assays and preformed biofilm biomass and viability assays. Scanning electron microscopy (SEM) was used to examine changes in cellular morphology. The results indicated that EGCG demonstrated activity against all isolates, with MICs ranging from 0.5 to 2 mg/mL, while AMP exhibited MIC values between 1.25 and 50 mg/mL. Importantly, the EGCG-AMP combination showed enhanced efficacy compared to either treatment alone, as indicated by a fractional inhibitory concentration index between 0.009 and 0.018. The most pronounced synergy was observed in 13 drug-resistant strains, where the MIC for EGCG dropped to 8 µg/mL (from 1 mg/mL alone) and that for AMP to 50 µg/mL (from 50 mg/mL alone), achieving a 125-fold and 1000-fold reduction, respectively. Time-kill assays revealed that the bactericidal effect of the EGCG-AMP combination occurred within 2 h. The mechanism of EGCG action includes the disruption of membrane permeability and biofilm eradication in a dose-dependent manner. SEM confirmed that the combination treatment consistently outperformed the individual treatments. This study underscores the potential of restoring AMP efficacy in combination with EGCG as a promising strategy for treating MDR E. coli infections.

Antifungal activity of dimethyl trisulfide and potential biocontrol against Alternaria alternata on postharvest Lycium barbarum

Abstract Lycium barbarum, is a medicine-food homology plant, so sustainable control of the postharvest Goji berry black mold caused by Alternaria alternata is particularly critical. In this paper, the impact of dimethyl trisulfide (DMTS) on A. alternata was studied, as well as the effects of DMTS on hypha ultrastructure, membrane permeability, reactive oxygen production, and essential enzymes of the cellular defense system. DMTS was shown to significantly inhibit the spore germination and hyphal growth of A. alternata in a concentration-dependent manner. The results of scanning electron microscopy (SEM) and transmission electron microscopy (TEM) showed that A. alternata cells were separated between cytoplasm and cell wall, organelles such as mitochondria were vacuolate or even disappeared, and the cytoplasm was fused. PI staining, conductivity measurements, and intracellular nucleic acid and protein content measurements showed that the permeability of the cell membrane changed after DTMS treatment, leading to the leakage of intracellular nucleic acid and protein. DMTS could also cause the accumulation of hydrogen peroxide (H2O2) and malondialdehyde (MDA), thus causing membrane oxidative damage. Through directly destroying the fungal cell and indirectly inducing membrane damage, DMTS can control the postharvest Goji berry black mold, and is a potential application prospect in the control of postharvest diseases.

Intracellular and Extracellular Metabolic Response of the Lactic Acid Bacterium Weissella confusa Under Salt Stress.

Weissella confusa is a member of the lactic acid bacterium group commonly found in many salt-fermented foods. Strains of W. confusa isolated from high-salinity environments have been shown to tolerate salt stress to some extent. However, the specific responses and mechanisms of W. confusa under salt stress are not fully understood. To study the effect of NaCl stress on W. confusa, growth performance and metabolite profiles of the strains were compared between a NaCl-free group and a 35% NaCl-treated group. Growth performance was assessed by measuring viable cell counts and examining the cells using scanning electron microscopy (SEM). Intracellular and extracellular metabolites were analyzed by non-targeted metabolomics based on liquid chromatography-mass spectrometry (LC-MS). It was found that the viable cell count of W. confusa decreased with increasing salinity, and cells could survive even in saturated saline (35%) medium for 24 h. When exposed to 35% NaCl, W. confusa cells exhibited surface pores and protein leakage. Based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, 42 different metabolites were identified in the cells and 18 different metabolites in the culture medium. These different metabolites were mainly involved in amino acid metabolism, carbohydrate metabolism, and nucleotide metabolism. In addition, salt-exposed cells exhibited higher levels of intracellular ectoine and lactose, whose precursors, such as aspartate, L-2,4-diaminobutanoate, and galactinol, were reduced in the culture medium. This study provides insight into the metabolic responses of W. confusa under salt stress, revealing its ability to maintain viability and alter metabolism in response to high NaCl concentrations. Key metabolites such as ectoine and lactose, as well as changes in amino acid and nucleotide metabolism, may contribute to its tolerance to salt. These findings may improve our understanding of the bacterium's survival mechanisms and have potential applications in food fermentation and biotechnology.

Amlodipine Ocular Delivery Restores Ferning Patterns and Reduces Intensity of Glycosylated Peak of Carrageenan-Induced Tear Fluid: An In-Silico Flexible Docking with IL-β1.

The tear ferning test can be an easy clinical procedure for the evaluation and characterization of the ocular tear film. The objective of this study was to examine the restoration of tear ferning patterns and reduction of glycosylation peak after amlodipine application in carrageenan-induced conjunctivitis. At the rabbit's upper palpebral region, carrageenan was injected for cytokine-mediated conjunctivitis. Ferning pattern and glycosylation of the tear fluid were characterized using various instrumental analyses. The effect of amlodipine was also examined after ocular instillation and flexible docking studies. Optical microscopy showed a disrupted ferning of the tear collected from the inflamed eye. FTIR of the induced tear fluid exhibited peaks within 1000-1200 cm-1, which might be due to the protein glycosylation absent in the normal tear spectrogram. The glycosylation peak reduced significantly in the tear sample collected from the amlodipine-treated group. Corresponding energy dispersive analysis showed the presence of sulphur, indicating protein leakage from the lacrimal gland in the induced group. The disappearance of sulphur from the treated group indicated its remedial effect. The flexible docking studies revealed a stronger binding mode of amlodipine with Interleukin-1β (IL-1β). The reduction in the intensity of the glycosylated peak and the restoration offering are probably due to suppression of IL-1β. This study may be helpful in obtaining primary information for drug discovery to be effective against IL-1β and proving tear fluid as a novel diagnostic biomarker.

Enzymatic Modulation of the Pulmonary Glycocalyx Enhances Susceptibility to Streptococcus pneumoniae.

The pulmonary epithelial glycocalyx is rich in glycosaminoglycans such as hyaluronan and heparan sulfate. Despite their presence, the importance of these glycosaminoglycans in bacterial lung infections remains elusive. To address this, we intranasally inoculated mice with Streptococcus pneumoniae in the presence or absence of enzymes targeting pulmonary hyaluronan and heparan sulfate, followed by characterization of subsequent disease pathology, pulmonary inflammation, and lung barrier dysfunction. Enzymatic degradation of hyaluronan and heparan sulfate exacerbated pneumonia in mice, as evidenced by increased disease scores and alveolar neutrophil recruitment. However, targeting epithelial hyaluronan in combination with S. pneumoniae infection further exacerbated systemic disease, indicated by elevated splenic bacterial load and plasma concentrations of proinflammatory cytokines. In contrast, enzymatic cleavage of heparan sulfate resulted in increased bronchoalveolar bacterial burden, lung damage, and pulmonary inflammation in mice infected with S. pneumoniae. Accordingly, heparinase-treated mice also exhibited disrupted lung barrier integrity as evidenced by higher alveolar edema scores and vascular protein leakage into the airways. This finding was corroborated in a human alveolus-on-a-chip platform, confirming that heparinase treatment also disrupts the human lung barrier during S. pneumoniae infection. Notably, enzymatic pretreatment with either hyaluronidase or heparinase also rendered human epithelial cells more sensitive to pneumococci-induced barrier disruption, as determined by transepithelial electrical resistance measurements, consistent with our findings in murine pneumonia. Taken together, these findings demonstrate the importance of intact hyaluronan and heparan sulfate in limiting pneumococci-induced damage, pulmonary inflammation, and epithelial barrier function and integrity.

Antibacterial activity of Azanza garckeana extracts (Malvaceae) in vitro and their potential use in respiratory infections

The ESKAPE pathogens, namely Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species, pose a significant threat to individuals with compromised immune system, including children, people with underlying illnesses and patients primarily infected with viruses. Significant mortality rates have been documented as a consequence of severe pneumonia resulting from bacterial respiratory tract infections. Azanza garckeana has been reported to possess antibacterial and anti-inflammatory activities. This study aimed to determine the antibacterial activity of A. garckeana leaf and bark extracts against P. aeruginosa, K. pneumoniae, A. baumannii, and S. aureus. The broth microdilution method was used to evaluate antibacterial activity. The most active extracts were subjected to phytochemical analysis to identify types of bioactive compounds present using gas chromatograph mass spectrometry (GC‒MS). The effect of the extracts on the integrity of the bacterial membrane was performed using nucleic acid and protein leakage assay. Acetone bark extract was assessed for its potential antibiofilm activity using K. pneumoniae. The toxicity profiling of the most potent extracts was performed using sheep erythrocytes and mouse peritoneal cells. The hexane bark extract exhibited greater potency by inhibiting the growth of S. aureus and A. baumannii at a concentration of 200 μg/mL. GC-MS identified the presence of important bioactive compounds including, β-carotene, 9-hexadecen-1-ol, (Z)-, hexadecanoic acid, methyl ester, and 2,4-di-tert-butylphenol. Acetone bark extract exhibited antibacterial activity through disruption of bacterial membrane integrity, observed through significant nucleic acid and protein leakage. The acetone bark extract displayed promising antibiofilm activity against K. pneumoniae. Importantly, the extracts showed minimal toxicity, demonstrating less than 30 % haemolytic activity in sheep erythrocytes and were not toxic to the mouse peritoneal cells, instead boosting their growth. These findings suggest that A. garckeana may serve as a potential source of antibacterial lead agents for the management of respiratory infections.

Solid- and Vapor-Phase Antibacterial Activities and Mechanisms of Essential Oils Against Fish Spoilage Bacteria.

Essential oils (EOs), regarded as secondary metabolites from plants, possess effective antibacterial properties. This study investigates the antibacterial efficacy of seven citrus EOs against six spoilage bacteria: Vibrio parahaemolyticus, V. harveyi, Photobacterium damselae, Shewanella putrefaciens, Carnobacterium divergens, and Lactobacillus pentosus. The antibacterial activity of these EOs was evaluated using solid- and vapor-phase applications. All tested EOs demonstrated effective antibacterial activity at a concentration of 294 μL/L against Gram-negative bacteria. Notably, lemon and orange EOs exhibited dose-dependent inhibition in both solid- and vapor-phase applications, with minimum effective concentrations ranging from 29.4 to 58.8 μL/L. Following treatment with lemon and orange EOs for 6 h at 1/4 minimum inhibitory concentration, leakage of intracellular DNA and proteins was observed, indicating damage to the cell membrane/wall. Proteomic analysis revealed distinct mechanisms: lemon EO impaired bacterial antioxidant defenses, while orange EO disrupted cell division, leading to reduced bacterial viability. These findings provide valuable insights into the potential of different EO application forms in controlling spoilage bacteria.

Curcuma longa rhizome extract: a potential antibiofilm agent against antibiotic-resistant foodborne pathogens

The traditional medicinal value of Curcuma longa (turmeric) and its potential relevance in modern healthcare suggests that traditional remedies and natural products can provide valuable solutions to contemporary challenges, such as combating biofilms and antibiotic-resistant pathogens, potentially offering new strategies for addressing health and safety issues in the fields of food and medicine. This study assessed the antibiofilm and antibacterial characterization of Curcuma longa rhizome extract against antibiotic-resistant foodborne pathogens. Gas Chromatography-Mass Spectrometry (GC-MS) and Fourier-transform infrared (FTIR) analysis were determined to check for the compounds, functional groups, and constituents of the plant extract. In-vitro antibiofilm and antibacterial bioassay of the extract were determined using standard bacteriological procedures. Potential mechanisms of the plant extract were also studied using standard biological methods. The important chemical constituents from the GC-MS extract of C. longa are arturmerone, cinnamyl angelate, tumerone, γ-atlantone, atlantone, α-atlantone, γ-atlantone and curlone. The FTIR analysis of the extract comprises alkyl halides, bromoalkanes, alkanes, ethylene molecules, arenes, amines, alcohols, sulfones, carboxylic acids and their derivatives, aromatic compounds, and phenols. The MIC of C. longa crude extract ranges from ethanol extract (0.03125 − 0.5 mg/mL) and acetone extract (0.0625 − 0.5 mg/mL). The MBC range is as follows: ethanol extract (0.125 − 1 mg/mL), acetone extract (0.125 − 1 mg/mL). The time-kill kinetics showed significant cell reduction with time. The bacterial isolates’ nucleic acids and protein leakage were consistent with increased extract concentration and time. There was a reduction in the biofilm cell on the shrimp surface and EPS with increased concentration and time. C. longa exerted significant anti-biofilm activity by removing existing biofilms, disrupting cell connections, and decreasing cells in biofilms. These findings can aid food protection from microbial contamination and prevent biofilms-related infections.

Serum protein and imaging biomarkers after intermittent steroid treatment in muscular dystrophy

Weekly Steroids in Muscular Dystrophy (WSiMD) was a pilot study to evaluate once weekly prednisone in patients with Limb Girdle and Becker muscular dystrophy (LGMD and BMD, respectively). At study endpoint, there were trends towards increased lean mass, reduced fat mass, reduced creatine kinase and improved motor function. The investigation was motivated by studies in mouse muscular dystrophy models in which once weekly glucocorticoid exposure enhanced muscle strength and reduced fibrosis. WSiMD participants provided blood samples for aptamer serum profiling at baseline and after 6 months of weekly steroids. A subset completed magnetic resonance (MR) evaluation of muscle at study onset and endpoint. At baseline compared to age and sex-matched healthy controls, the aggregate serum protein profile in the WSiMD cohort was dominated by muscle proteins, reflecting leak of muscle proteins into serum. Disease status produced more proteins differentially present in serum compared to steroid-treatment effect. Nonetheless, a response to prednisone was discernable in the WSiMD cohort, even at this low dose. Glucocorticoids decreased muscle proteins and increased certain immune process- and matrix-associated proteins. Muscle MR fat fraction showed trends with functional status. The prednisone-responsive markers could be used in larger trial of prednisone efficacy.

Response mechanism of Saccharomyces cerevisiae under benzoic acid stress in ethanol fermentation

Sugarcane molasses is an ideal economical raw material for ethanol production because of its wide availability, low cost and nutrient content. However, benzoic acid compounds with toxic effects on yeast cells are commonly found in sugarcane molasses. At present, the molecular mechanism of the toxic effects of benzoic acid on Saccharomyces cerevisiae has not been elucidated. Here, the toxic effect of exogenous benzoic acid on S. cerevisiae GJ2008 cells was studied, and the genes differentially expressed in S. cerevisiae GJ2008 after 1.2 g/L benzoic acid stress were identified via Illumina RNA-Seq technology. The results indicated that benzoic acid significantly inhibited yeast cell growth, prolonged their rapid growth period, and ultimately reduced their biomass. During ethanol fermentation using 250 g/L sucrose under 1.2 g/L benzoic acid stress, several adverse effects were observed, such as high residual sugar content, low ethanol concentration and low fermentation efficiency. In addition, the cell morphology was damaged, the cell membrane permeability increased, intracellular nucleic acid and protein leakage increased, and the malondialdehyde content significantly increased. Moreover, the cells protected themselves by significantly increasing the intracellular glycerol content. Fourier transform infrared spectroscopy proved that benzoic acid could reduce the degree of unsaturation and increase cell membrane permeability by changing the yeast cell wall and cell membrane composition, leading to cell damage and even death. Transcriptomic analysis revealed that under benzoic acid stress, the expression of genes associated with sucrose and starch metabolism, thiamine metabolism, the glycolysis pathway, fructose and mannose metabolism, galactose metabolism and ABC transporters was significantly downregulated. The expression of genes related to ribosomes, lipid metabolism, ribosome biosynthesis, nucleic acid metabolism, arginine and proline metabolism, RNA polymerase, metabolism related to cofactor synthesis, and biosynthesis of valine, leucine, and isoleucine was significantly upregulated. These results indicated that benzoic acid inhibited glycolysis and reduced sugar absorption and utilization and ATP energy supply in yeast cells. In response to stress, genes related to the ribosome bioanabolic pathway were upregulated to promote protein synthesis. On the other hand, the expression of ELO1, SUR4, FEN1 and ERG1 was upregulated, which led to extension of long-chain fatty acids and accumulation of ergosterol to maintain cell membrane structure. In conclusion, this paper provides important insights into the mechanism underlying the toxicity of benzoic acid to yeast cells and for realizing high-concentration ethanol production by sugarcane molasses fermentation.

Molecular Mechanism: Inhibition of Fusarium oxysporum T-2 Toxin Synthesis by Surfactin in Dried Fish: Induction of Yap1 Nucleation by ROS Accumulation.

(1) T-2 toxin synthesized by Fusarium oxysporum (F. oxysporum) can cause deterioration of dried fish and endanger human health. (2) The molecular mechanism by which antibacterial lipopeptides surfactin inhibited F. oxysporum growth and toxin production was elucidated by investigating the intracellular ROS production pathway and the subcellular distribution and transcriptional activity of the transcription factor Yap1 and its regulation of Tri5 gene in F. oxysporum. (3) Surfactin caused hyphal damage and nucleic acid and protein leakage; thus, the growth of F. oxysporum was disrupted. Surfactin's excessive accumulation of intracellular ROS triggered the translocation of transcription factor Yap1 into the nucleus, resulting in toxin cluster gene Tri5 expression inhibition, thereby blocking T-2 toxin synthesis. (4) This is a novel mechanism by which surfactin inhibits the growth and T-2 toxin synthesis of F. oxysporum from multiple aspects, including cell structural integrity and the ROS-Yap1 signaling pathway. (5) This study provides a theoretical basis for the application of surfactin in the antifungal control of aquatic dry products.