Oxidase Production Research Articles

Anti-inflammatory Fatty Acid Derivatives From Mangrove-derived Actinomycetes Streptomyces sp.

Mangrove-derived actinomycetes are prolific chemical sources of complex and novel natural products, providing an excellent chance for new drug discovery. The chemical investigation of the mangrove-derived Streptomycessundarbansensis 06037, led to the discovery of two previously undescribed enone fatty acids (1-2), one new phenylpropionate derivate (3), along with the isolation of the ten known components (4-13). Those chemical structures of isolates were elucidated on the basis of the analysis of diverse spectroscopic data. Initial anti-inflammatory tests of 1-3 in lipopolysaccharide (LPS)-activated RAW 264.7 murine macrophage cells revealed that compound 1 possess significant inhibitory effect on the production of Nitro oxidase (NO), with the IC50 value around 15.33 ± 1.32 μM, together with the suppression of NF-κB phosphorylation and reducing the release of oxygen species (ROS) in RAW 264.7 macrophages, those results indicated that compound 1 may exert its anti-inflammatory activity through a reduction in ROS level and the suppression of NF-κB activation pathway.

Para-coumaric acid and cinnamic acid enhance resistance of Agaricus bisporus mushrooms to Brown blotch disease caused by Pseudomonas tolaasii

The effects of postharvest pre-treatments with para-coumaric acid (PCA) and cinnamic acid (CA) against brown blotch disease in Agaricus bisporus mushrooms caused by Pseudomonas tolaasii were investigated. PCA and CA at the concentration of 300 mg L−1 provided the highest inhibition ratio of 28.9% and 20.9%, respectively, on the in vitro growth of P. tolaasii. In addition, PCA and CA pre-treatments dramatically reduced the P. tolaasii-caused brown blotch disease in postharvest A. bisporus. Biochemical analysis revealed that PCA and CA inhibited P. tolaasii-induced activation of polyphenol oxidase and melanin production, increased activities of β-N-acetylglucosaminidase, β-1,3-glucanase and chitinase, promoted phenylpropanoid biosynthetic enzyme activities and metabolite production, enhanced antioxidant enzyme activities, and decreased malondialdehyde content in A. bisporus mushrooms. Collectively, these results demonstrated that PCA and CA can induce resistance to postharvest brown blotch disease in A. bisporus mushrooms, and have good prospects for disease control in edible mushrooms.

Multiple levels of transcriptional regulation control glycolate metabolism in Paracoccus denitrificans.

The hydroxyacid glycolate is a highly abundant carbon source in the environment. Glycolate is produced by unicellular photosynthetic organisms and excreted at petagram scales to the environment, where it serves as growth substrate for heterotrophic bacteria. In microbial metabolism, glycolate is first oxidized to glyoxylate by the enzyme glycolate oxidase. The recently described β-hydroxyaspartate cycle (BHAC) subsequently mediates the carbon-neutral assimilation of glyoxylate into central metabolism in ubiquitous Alpha- and Gammaproteobacteria. Although the reaction sequence of the BHAC was elucidated in Paracoccus denitrificans, little is known about the regulation of glycolate and glyoxylate assimilation in this relevant alphaproteobacterial model organism. Here, we show that regulation of glycolate metabolism in P. denitrificans is surprisingly complex, involving two regulators, the IclR-type transcription factor BhcR that acts as an activator for the BHAC gene cluster, and the GntR-type transcriptional regulator GlcR, a previously unidentified repressor that controls the production of glycolate oxidase. Furthermore, an additional layer of regulation is exerted at the global level, which involves the transcriptional regulator CceR that controls the switch between glycolysis and gluconeogenesis in P. denitrificans. Together, these regulators control glycolate metabolism in P. denitrificans, allowing the organism to assimilate glycolate together with other carbon substrates in a simultaneous fashion, rather than sequentially. Our results show that the metabolic network of Alphaproteobacteria shows a high degree of flexibility to react to the availability of multiple substrates in the environment.IMPORTANCEAlgae perform ca. 50% of the photosynthetic carbon dioxide fixation on our planet. In the process, they release the two-carbon molecule glycolate. Due to the abundance of algae, massive amounts of glycolate are released. Therefore, this molecule is available as a source of carbon for bacteria in the environment. Here, we describe the regulation of glycolate metabolism in the model organism Paracoccus denitrificans. This bacterium uses the recently characterized β-hydroxyaspartate cycle to assimilate glycolate in a carbon- and energy-efficient manner. We found that glycolate assimilation is dynamically controlled by three different transcriptional regulators: GlcR, BhcR, and CceR. This allows P. denitrificans to assimilate glycolate together with other carbon substrates in a simultaneous fashion. Overall, this flexible and multi-layered regulation of glycolate metabolism in P. denitrificans represents a resource-efficient strategy to make optimal use of this globally abundant molecule under fluctuating environmental conditions.

NAMPT enhances LOX expression and promotes metastasis in human chondrosarcoma cells by inhibiting miR-26b-5p synthesis.

Chondrosarcoma is a malignant bone tumor that emerges from abnormalities in cartilaginous tissue and is related with lung metastases. Nicotinamide phosphoribosyltransferase (NAMPT) is an adipocytokine reported to enhance tumor metastasis. Our results from clinical samples and the Gene Expression Omnibus data set reveal that NAMPT levels are markedly higher in chondrosarcoma patients than in normal individuals. NAMPT stimulation significantly increased lysyl oxidase (LOX) production in chondrosarcoma cells. Additionally, NAMPT increased LOX-dependent cell migration and invasion in chondrosarcoma by suppressing miR-26b-5p generation through the c-Src and Akt signaling pathways. Overexpression of NAMPT promoted chondrosarcoma metastasis to the lung in vivo. Furthermore, knockdown of LOX counteracted NAMPT-facilitated metastasis. Thus, the NAMPT/LOX axis presents a novel target for treating the metastasis of chondrosarcoma.

#944 Soluble uric acid inhibits neutrophil functions during infections in patients with kidney disease

Abstract Background and Aims Chronic kidney disease (CKD) is one of the most common diseases affecting 8-16% of the world population. Beside cardiovascular complications, infections including sepsis are the second most common cause of death in patients with acute kidney injury (AKI) and CKD. However, the underlying pathomechanisms that contribute to the secondary immunodeficiency related to kidney disease [1] are not well understood. Recent evidence suggests that CKD-related hyperuricemia characterised by elevated serum uric acid (UA) levels suppresses immune cell functions during sterile inflammation [2]. Whether this is also the case during host defense is subject of our current investigation. In this study, we hypothesized that soluble UA inhibits neutrophil effector functions and therefore contributes to the increased infection risk in patients with kidney disease. Method Blood neutrophils were isolated from healthy individuals as well as from patients with kidney diseases, and incubated ex vivo in the presence or absence of 10 mg/dl soluble UA prior to stimulation with bacterial peptides including LPS. Immune cell functions including cytoskeletal changes, immune cell activation, maturation, endo- and phagocytosis (pHrodo Dextran particles, IgG-FITC beads, pHrodo E. coli bioparticles), ROS production, and neutrophil extracellular trap (NET) formation were analysed and quantified using flow cytometry, fluorescence and colorimetric assays, and fluorescence microscopy. In addition, to verify our results we stimulated neutrophils from healthy individuals with sera from hyperuricemic CKD patients. Results Our results show for the first time that soluble UA significantly inhibits the ability of neutrophils to endocytose small particles and phagocytose beads in neutrophils from healthy individuals comparable to the inhibitory effect of Cytochalasin D, an inhibitor of endo- and phagocytosis. Interestingly, unlike neutrophils from AKI patients, neutrophils from CKD patients were significantly less able to phagocytose beads compared to healthy controls with and without stimulation. To mimic the uptake of pathogens more physiologically, pHrodo E. coli bioparticles were used. We found that soluble UA inhibited the phagocytosis of such bioparticles in neutrophils. In addition, our investigation into the formation of ROS showed that soluble UA inhibited specifically nitric oxide, peroxynitrite, hydroxyl radicals and hydrogen peroxide, but to a lower extent the formation of superoxides in neutrophils from healthy individuals, similar to the effectiveness of the radical scavenger N-acetylcysteine. These data were comparable with our cohort study of neutrophils from CKD patients. To examine the mechanism of reduced ROS production, we inhibited NADPH oxidase (DPI) and MPO (4-ABAH) in neutrophils and found that soluble UA has similar inhibitory effects on ROS production like DPI whereas 4-ABAH was more potent. This indicated that soluble UA modulates NADPH oxidase and ROS production, which subsequently resulted in reduced NET formation in neutrophils. Conclusion Our data identify soluble UA as potential immune regulator of the secondary immunodeficiency in patients with kidney disease by inhibiting the endo- and phagocytosis of particles, NADPH-mediated ROS production and NET formation in neutrophils, processes that are important to kill pathogens and fight an infection. Thus, specifically targeting UA with urate-lowering therapy might overcome the suppressed host defence against infection in patients with kidney disease.

Lysyl Oxidase Production by Murine C3H10T1/2 Mesenchymal Stem Cells Is Increased by TGFβs and Differentially Modulated by Mechanical Stimuli.

Tendons are frequently injured and have limited regenerative capacity. This motivates tissue engineering efforts aimed at restoring tendon function through strategies to direct functional tendon formation. Generation of a crosslinked collagen matrix is paramount to forming mechanically functional tendon. However, it is unknown how lysyl oxidase (LOX), the primary mediator of enzymatic collagen crosslinking, is regulated by stem cells. This study investigates how multiple factors previously identified to promote tendon formation and healing (transforming growth factor [TGF]β1 and TGFβ2, mechanical stimuli, and hypoxia-inducible factor [HIF]-1α) regulate LOX production in the murine C3H10T1/2 mesenchymal stem cell (MSC) line. We hypothesized that TGFβ signaling promotes LOX activity in C3H10T1/2 MSCs, which is regulated by both mechanical stimuli and HIF-1α activation. TGFβ1 and TGFβ2 increased LOX levels as a function of concentration and time. Inhibiting the TGFβ type I receptor (TGFβRI) decreased TGFβ2-induced LOX production by C3H10T1/2 MSCs. Low (5 mPa) and high (150 mPa) magnitudes of fluid shear stress were applied to test impacts of mechanical stimuli, but without TGFβ2, loading alone did not alter LOX levels. Low loading (5 mPa) with TGFβ2 increased LOX at 7 days greater than TGFβ2 treatment alone. Neither HIF-1α knockdown (siRNA) nor activation (CoCl2) affected LOX levels. Ultimately, results suggest that TGFβ2 and appropriate loading magnitudes contribute to LOX production by C3H10T1/2 MSCs. Potential application of these findings includes treatment with TGFβ2 and appropriate mechanical stimuli to modulate LOX production by stem cells to ultimately control collagen matrix stiffening and support functional tendon formation.

Effects of exogenous autoinducer-2 precursor on the ability of Lactiplantibacillus plantarum SH7 to degrade biogenic amines: In vitro and in a dry-sausage model

The effects of the exogenous autoinducer-2 (AI-2) precursor, 4,5-dihydroxy-2,3-pentanedione (DPD; 0, 300, 600, and 900 nmol/L), on the degradation capacity of Lactiplantibacillus plantarum SH7 toward biogenic amines (BA) were investigated both in vitro and in a dry-sausage model. L. plantarum SH7 exhibited a significant increase in BA-degradation capacity (putrescine, cadaverine, and histamine) under DPD treatments compared to the absence of DPD treatment in vitro. Additionally, a significant increase in the production of AI-2 signal molecule and diamine oxidase in L. plantarum SH7 was observed with the addition of exogenous DPD (p < 0.05). In the dry-sausage model, the contents of BAs (tryptamine, 2-phenethylamine, putrescine, cadaverine, and histamine) significantly decreased with the addition of DPD (p < 0.05). Meanwhile, the sausage with 600-nM DPD exhibited the lowest cadaverine content (11.13 mg/kg), while the sausage with 900-nM DPD exhibited the lowest tryptamine, putrescine, and histamine contents (12.44, 11.15, and 13.21 mg/kg, respectively). Results suggest that an optimal concentration of exogenous DPD can enhance the capacity of L. plantarum SH7 to degrade BAs and that the degradation of BAs by L. plantarum SH7 may be dependent on the LuxS/AI-2 quorum sensing (QS) system. This study provides the theoretical basis for the development of functional starter cultures of lactic acid bacteria with BA-degradation ability based on targeted LuxS/AI-2 QS system regulation and reducing the accumulation of BAs in fermented meat products.

Producción de oxidasas por Trametes versicolor crecido sobre residuos verdes y sobre espuma de poliuretano en fermentación en estado sólido: Un estudio comparativo

Green waste (GW) is generated by the maintenance of public or private green spaces. It is necessary to find ecological alternatives for GW utilization, aiming to avoid accumulation of this material at the environment. In this research, the production of laccase (Lac), lignin peroxidase (LiP), manganese peroxidase (MnP) and unspecific peroxygenase (UnP) produced by Trametes versicolor grown on GW as a substrate and on polyurethane foam (PUF) as an inert support in solid state fermentation was evaluated. T. versicolor showed higher values of Lac, MnP, UnP and LiP activities (34, 943, 1023 and 766 U/gS, respectively) when grown on GW than when grown on PUF (10.9, 588, 559 and 229 U/gS, respectively). These results suggest that T. versicolor produced Lac inducible and constitutively, while LiP, MnP and UnP were induced by GW at the beginning of fungal growth, however, these enzymes were constitutive and inducible during the rest of the fermentation. The production of oxidases and peroxidases was induced and increased by GW. It is suggested that LiP is involved (as a constitutive enzyme) at the beginning of the exponential phase, while MnP and UnP participate in fungal growth at the end of fermentation. To our knowledge, this is the first detailed study on the main lignincellulose-degrading fungal enzymes involved in GW degradation by fungi. In particular, the relevance of UnP was showed as peroxidase involved in lignocellulosic substrates biodegradation.

Comparative Effect of Seed Coating and Biopriming of Bacillus aryabhattai Z-48 on Seedling Growth, Growth Promotion, and Suppression of Fusarium Wilt Disease of Tomato Plants.

Beneficial plant microbes can enhance the growth and quality of field crops. However, the benefits of microbes using cheap and efficient inoculation methods are still uncommon. Seed coating with biocontrol agents can reduce the amount of inocula along with having the potential for large-scale application. Hence, in this research work, the comparative potential of tomato seed coating and biopriming with Bacillus aryabhattai Z-48, harboring multiple plant-beneficial traits, to suppress Fusarium wilt disease along with its beneficial effect on seedling and plant growth promotion was analyzed. Among two bacterial strains, B. aryabhattai Z-48 was able to antagonize the mycelial growth of Fusarium oxysporum f.sp. lycopersici in vitro and its application as a seed coating superiorly benefited seedling traits like the germination percentage, vigor index, and seedling growth index along with a reduced germination time. The seed coating with B. aryabhattai Z-48 resulted in significant increases in the shoot length, root length, dry biomass, and total chlorophyll contents when compared with the bioprimed seeds with the same bacterial strain and non-inoculated control plants. The seed coating with B. aryabhattai Z-48 significantly reduced the disease index (>60%) compared with the pathogen control during pot trials. Additionally, the seed coating with B. aryabhattai Z-48 resulted in a significantly higher production of total phenolics, peroxidase, polyphenol oxidase, and phenylalanine ammonia lyase enzyme in tomato plants. The GC/MS-based non-targeted metabolic profiling indicated that the seed coating with B. aryabhattai Z-48 could cause large-scale metabolite perturbations in sugars, sugar alcohols, amino acids, and organic acids to increase the fitness of tomato plants against biotic stress. Our study indicates that a tomato seed coating with B. aryabhattai Z-48 can improve tomato growth and suppress Fusarium wilt disease effectively under conventional agricultural systems.

Genome analysis of vB_SupP_AX, a novel N4-like phage infecting Sulfitobacter.

Sulfitobacter is a bacterium recognized for its production of AMP-independent sulfite oxidase, which is instrumental in the creation of sulfite biosensors. This capability underscores its ecological and economic relevance. In this study, we present a newly discovered phage, Sulfitobacter phage vB_SupP_AX, which was isolated from Maidao of Qingdao, China. The vB_SupP_AX genome is linear and double-stranded and measures 75,445 bp with a GC content of 49%. It encompasses four transfer RNA (tRNA) sequences and 79 open reading frames (ORFs), one of which is an auxiliary metabolic gene encoding thioredoxin. Consistent with other N4-like phages, vB_SupP_AX possesses three distinct RNA polymerases and is characterized by the presence of four tRNA molecules. Comparative genomic and phylogenetic analyses position vB_SupP_AX and three other viral genomes from the Integrated Microbial Genomes/Virus v4 database within the Rhodovirinae virus subfamily. The identification of vB_SupP_AX enhances our understanding of virus-host interactions within marine ecosystems.

The Effect of Soursop Leaf Extract in Reducing Pain in Gout Arthritis

Gout arthritis is a disease characterized by pain caused by deposits of monosodium crystals that accumulate in the joints due to high levels of uric acid in the blood. Currently, pain management treatment for gout arthritis patients focuses on pharmacological treatment by providing analgesics to relieve pain. Soursop leaf extract has been proven to have an effect in reducing pain, but there has been no research regarding the effect of boiled soursop leaves to reduce it. The aim of this study was to analyze the effect of giving soursop leaf decoction on reducing pain in gout arthritis patients. The research used a quasi-experimental design with a one group pre-posttest design. The sample selected by using purposive sampling technique, totaling 18 people. The independent variable was soursop leaf decoction. The dependent variable was the pain in gout arthritis. The data analysis used the Wilcoxon Signed Ranks Test statistical test. The compounds contained in soursop leaves function as a strong analgesic (pain reliever) and have antioxidant properties. The antioxidant properties contained in soursop leaves can reduce the formation of uric acid by inhibiting the production of the enzyme xanthine oxidase. This combination of analgesic and anti-inflammatory properties can reduce gout. This study found that there was an effect of giving soursop leaf decoction on reducing pain in gout arthritis patients.

Ecological versatility and biotechnological promise: Comprehensive characterization of the isolated thermophilic Bacillus strains.

This study focuses on isolated thermophilic Bacillus species' adaptability and physiological diversity, highlighting their ecological roles and potential industrial applications. We specifically investigated their capacity to thrive in extreme conditions by examining their environmental tolerances and adaptations at the metabolic and genetic levels. The primary objective is to evaluate the suitability of these species for biotechnological applications, considering their resilience in harsh environments. We conducted a comparative analysis of the environmental adaptability parameters for various Bacillus species. This included examining growth temperature ranges, pH tolerance, oxygen requirements, carbohydrate fermentation patterns, colony morphology, enzymatic activities, and genetic properties. Controlled laboratory experiments provided the data, which were then analyzed to determine patterns of adaptability and diversity. The research revealed that Bacillus species could endure temperatures as high as 73°C, with a generally lower growth limit at 43°C. However, strains TBS35 and TBS40 were exceptions, growing at 37°C. Most strains preferred slightly alkaline conditions (optimal pH 8), but TBS34, TBS35, and TBS40 exhibited adaptations to highly alkaline environments (pH 11). Oxygen requirement tests classified the species into aerobic, anaerobic, and facultative aerobic categories. Genetic analysis highlighted variations in DNA concentrations, 16s rRNA gene lengths, and G+C content across species. Although glucose was the primary substrate for carbohydrate fermentation, exceptions indicated metabolic flexibility. The enzymatic profiles varied, with a universal absence of urease and differences in catalase and oxidase production. Our findings underscore thermophilic Bacillus species' significant adaptability and diversity under various environmental conditions. Their resilience to extreme temperatures, pH levels, varied oxygen conditions, and diverse metabolic and genetic features emphasize their potential for biotechnological applications. These insights deepen our understanding of these species' ecological roles and highlight their potential industrial and environmental applications.

HISTORICAL HERITAGE IN ANTARCTICA, THE CASA MONETA MUSEUM: A FIRST APPROACH TO THE CHARACTERIZATION OF XYLOPHAGOUS FUNGI AND THEIR POTENTIAL ROLE IN WOOD DAMAGE

The Casa Moneta Museum is located at the base located at an isthmus located on Laurie Island in the South Orkney Islands archipelago. Currently, the museum shows signs of deterioration of the wood (discoloration, fibrous appearance and presence of mycelium) and it is necessary to characterize this mycobiota to analyze the potential role in the biodegradation process. An assessment of the potential risk associated with fungal communities could provide valuable information for taking conservation action at the Casa Moneta Museum. The aim of this work was to evaluate the extracellular oxidase production of fungi recovered from deteriorated wood at the museum and to analyze the biodegradation risk index of three sampled sites in the building. The enzymatic profile showed the potential degradation of Cadophora spp. and Tulasnella albida strains. The analysis performed integrated data on several parameters like occurrences, species diversity, dominant species, wind direction, relative humidity, growth temperature, type of dispersion propagules and substrate, indicating Site 2 has a major biodegradation risk index which suggests that this site should be the immediate target for conservation efforts.

Hypersecretory production of glucose oxidase in Pichia pastoris through combinatorial engineering of protein properties, synthesis, and secretion.

Glucose oxidase (EC 1.1.3.4, GOD) is a widely used industrial enzyme. To construct a GOD-hyperproducing Pichia pastoris strain, combinatorial strategies have been applied to improve GOD activity, synthesis, and secretion. First, wild-type GOD was subjected to saturation mutagenesis to obtain an improved variant, MGOD1 (V20W/T30S), with 1.7-fold higher kcat /KM . Subsequently, efficient signal peptides were screened, and the copy number of MGOD1 was optimized to generate a high-producing strain, 8GM1, containing eight copies of AOX1 promoter-GAS1 signal peptide-MGOD1 expression cassette. Finally, the vesicle trafficking of 8GM1 was engineered to obtain the hyperproducing strain G1EeSe co-expressing the trafficking components EES and SEC. 22, and the EES gene (PAS_chr3_0685) was found to facilitate both protein secretion and production for the first time. Using these strategies, GOD secretion was enhanced 65.2-fold. In the 5-L bioreactor, conventional fed-batch fermentation without any process optimization resulted in up to 7223.0 U/mL extracellular GOD activity (3.3-fold higher than the highest level reported to date), with almost only GOD in the fermentation supernatant at a protein concentration of 30.7 g/L. Therefore, a GOD hyperproducing strain for industrial applications was developed, and this successful case can provide a valuable reference for the construction of high-producing strains for other industrial enzymes.

Improved production of Bacillus subtilis cholesterol oxidase by optimization of process parameters using response surface methodology

BackgroundCholesterol oxidase has numerous biomedical and industrial applications. In the current study, a new bacterial strain was isolated from sewage and was selected for its high potency for cholesterol degradation (%) and production of high cholesterol oxidase activity (U/OD600). ResultsBased on the sequence of 16S rRNA gene, the bacterium was identified as Bacillus subtilis. The fermentation conditions affecting cholesterol degradation (%) and the activity of cholesterol oxidase (U/OD600) of B. subtilis were optimized through fractional factorial design (FFD) and response surface methodology (RSM). According to this sequential optimization approach, 80.152% cholesterol degradation was achieved by setting the concentrations of cholesterol, inoculum size, and magnesium sulphate at 0.05 g/l, 6%, and 0.05 g/l, respectively. Moreover, 85.461 U of cholesterol oxidase/OD600 were attained by adjusting the fermentation conditions at initial pH, 6; volume of the fermentation medium, 15 ml/flask; and concentration of cholesterol, 0.05 g/l. The optimization process improved cholesterol degradation (%) and the activity of cholesterol oxidase (U/OD600) by 139% and 154%, respectively. No cholesterol was detected in the spectroscopic analysis of the optimized fermented medium via gas chromatography-mass spectroscopy (GC–MS). ConclusionThe current study provides principal information for the development of efficient production of cholesterol oxidase by B. subtilis that could be used in various applications.

Panton-Valentine leukocidin-induced neutrophil extracellular traps lack antimicrobial activity and are readily induced in patients with recurrent PVL + -Staphylococcus aureus infections.

Staphylococcus aureus strains that produce the toxin Panton-Valentine leukocidin (PVL-SA) frequently cause recurrent skin and soft tissue infections. PVL binds to and kills human neutrophils, resulting in the formation of neutrophil extracellular traps (NETs), but the pathomechanism has not been extensively studied. Furthermore, it is unclear why some individuals colonized with PVL-SA experience recurring infections whereas others are asymptomatic. We thus aimed to (1) investigate how PVL exerts its pathogenicity on neutrophils and (2) identify factors that could help to explain the predisposition of patients with recurring infections. We provide genetic and pharmacological evidence that PVL-induced NET formation is independent of NADPH oxidase and reactive oxygen species production. Moreover, through NET proteome analysis we identified that the protein content of PVL-induced NETs is different from NETs induced by mitogen or the microbial toxin nigericin. The abundance of the proteins cathelicidin (CAMP), elastase (NE), and proteinase 3 (PRTN3) was lower on PVL-induced NETs, and as such they were unable to kill S. aureus. Furthermore, we found that neutrophils from affected patients express higher levels of CD45, one of the PVL receptors, and are more susceptible to be killed at a low PVL concentration than control neutrophils. Neutrophils from patients that experience recurring PVL-positive infections may thus be more sensitive to PVL-induced NET formation, which might impair their ability to combat the infection.

Characterization and immobilization of extracellular broad pH active multicopper oxidase from Streptomyces flavomacrosporus for dye-discoloration

The toxic and recalcitrant dyes negatively impact our environment. The biological breakdowns of these toxic compounds using laccases have been explored to degrade these dyes effectively. Despite the low redox potential, compared to fungal laccases, the ability of Streptomyces laccases to work under alkaline pH against a wide range of substrates and unusual resistance to inhibitors opens up new opportunities for their commercial applications. The present study evaluated Streptomyces isolates for multicopper oxidase (MCO) production using 2,6-dimethoxyphenol (DMP), syringaldazine and guaiacol as substrates. The biochemical characterisation of purified laccase enzyme showed that the enzyme is active between 28 and 50 °C and has a pH range of 5–8, depending upon the substrates. The immobilization of the laccase showed that the beads retained their size under acidic conditions but did not exhibit any enzyme activity. The immobilized enzyme exhibited maximum activity under alkaline conditions. The alginate laccase immobilized on beads can be reused two to three times without any significant loss of activity. The potential of free and immobilized MCO from Streptomyces significantly reduced the absorption spectra of different dyes, indicating its role in dye-discoloration, which can be used for various industrial processes and industrial effluent treatment.

Methanol Formation, Toxicity and its Impact on the Human Nervous System and Liver

Methanol poisoning is a severe public health issue that causes disease and death, particularly in developing countries. Exposure can occur intentionally or unintentionally through several industrial products such as cleansers, solvents, antifreeze, perfumes, and paints. The toxic dose of methanol is approximately 1g/kg, emphasizing its potent toxicity. Methanol intoxication is chiefly caused by the harmful metabolites which are formed during its breakdown. These metabolites encourage metabolic acidosis. Formic acid, a methanol breakdown product, troubles mitochondria, distressing cytochrome oxidase and ATP production. Methanol-related problems include neurological disorders, vision problems, liver impact, Alzheimer’s, and Parkinson's. Key lab tests, including osmolality and anionic gap, aid in recognizing alcohol harm. Furthermore, detecting a faint smell of methanol in the breath, observing visual disturbances, and altered sensory neurons, seeing a history of abdominal pain, and conducting routine checkups support the early diagnosis of methanol poisoning. Treatment using prompt gastric lavage, sodium bicarbonate given intravenously, management with ethanol, folate-dependent folate systems suggest, dialysis, competent ADH inhibitor “pyrazole”, specific antidote “Fomepizole” can be used to treat methanol poisoning. Regular toxicologist check-ups are critical in such cases. Definite symptoms like basal ganglia issues and eye problems require consultations with neurologists and ophthalmologists. For cases needing hemodialysis or liver tissue recovery, consulting a nephrologist is essential. It is suggested to immediately act to prevent harm, especially for industrial workers, rural alcohol sellers, those with alcoholism, and vulnerable youth vulnerable to methanol poisoning.

CRISPR/Cas9 System-Mediated Multi-copy Expression of an Alkaline Serine Protease in Aspergillus niger for the Production of XOD-Inhibitory Peptides.

CRISPR/Cas9 system-mediated multi-copy expression of an alkaline serine protease (AoproS8) from Aspergillus oryzae was successfully built in Aspergillus niger. Furthermore, AoproS8 was continuously knocked in the glaA, amyA, and aamy gene loci in A. niger to construct multi-copy expression strains. The yield of the AoproS8 3.0 strain was 2.1 times higher than that of the AoproS8 1.0 strain. Then, a high protease activity of 11,023.2 U/mL with a protein concentration of 10.8 mg/mL was obtained through fed-batch fermentation in a 5 L fermenter. This is the first report on the high-level expression of alkaline serine proteases in A. niger. AoproS8 showed optimal activity at pH 9.0 and 40 °C. It was used for the production of xanthine oxidase (XOD)-inhibitory peptides from eight food processing protein by-products. Among them, the duck hemoglobin hydrolysates showed the highest XOD-inhibitory activity with an IC50 value of 2.39 mg/mL. Thus, our work provides a useful way for efficient expression of proteases in A. niger and high-value utilization of protein by-products.

Biocontrol adeptness of bacterial endophytes antagonistic to Colletotrichum spp. causing Colletotrichum leaf disease in rubber (Hevea brasiliensis) and harnessing its plant growth-promoting traits

Colletotrichum Leaf Disease (CLD) is one of the major leaf diseases which significantly affect growth and yield of natural rubber (NR). Development of eco-friendly disease management strategies is the need of the hour as various commercial fungicides are banned due to their detrimental effect on environment. The present study characterized antagonistic activities and plant growth promoting properties of five bacterial endophytes, isolated from rubber leaves and roots, against Colletotrichum acutatum and C. gloeosporioides, causing CLD in rubber. In the dual culture test, Bacillus sp. and Stenotrophomonas sp. showed 62 and 60 per cent inhibition of radial growth (PIRG). With the culture filtrate, Stenotrophomonas sp. (RLR1), Bacillus sp. (RRF1), B. subtilis (RLF1), Ochrobactrum anthropi (RLF3), and Burkholderia lata (RLF15) recorded suppression of growth up to 100%. These endophytic bacterial strains were identified based on biochemical characteristics and through 16S rRNA gene sequencing. The isolates were also tested for twenty five plant growth promoting traits. All five isolates responded positively for cellulase production, siderophore production, citrate utilization, and oxidase production. However, all five strains responded negatively to chitinase, urease, phosphate solubilisation, hydrogen sulphide production, and fluorescent pigment production. These results confirmed that the five identified bacterial isolates possessing growth-promoting characteristics have the potential to be used as biocontrol agents against Colletotrichum spp., a major devastating pathogen of rubber.