Search Results (47)

Botryosphaeria dothidea is the causative agent of Chinese hickory trunk canker, which poses significant threat to the production of Chinese hickory (Carya cathayensis Sarg.). Previous studies reported that endophytic–pathogenic phase transition, also referred to as latent infection, plays an important role in the interaction of Botryosphaeria dothidea with various host plants, including Chinese hickory. However, the mechanism underlying this phase transition is not well understood. Here, we employed RNA-Seq to investigate transcriptional changes in B. dothidea during its phase transition upon interaction with Chinese hickory. A co-expression network was generated based on 6391 differentially expressed genes (DEGs) identified from different infection stages and temperature treatments. One co-expressed module was found that highly correlated with temperature treatments which simulated conditions of B. dothidea latent infection in the field. Subsequently, 53 hub genes were detected, and gene ontology (GO) enrichment analysis revealed three categories of enriched GO terms: transmembrane transport or activity, ion homeostasis or transport, and carbohydrate metabolism. One PacC transcriptional factor (BDLA_00001555, an ambient pH regulator), and one endo-β-1,3-glucanase (BDLA_00010249) were specifically upregulated under temperature treatments that corresponded with the activation stage of B. dothidea’s pathogenic state. The knockout mutant strain of BDLA_00001555 demonstrated defective capability upon the activation of the pathogenic state. This confirmed that BDLA_00001555, the PacC transcriptional factor, plays an important role in the latent infection phase of B. dothidea. Our findings provide insights into the pathogenic mechanism of Chinese hickory trunk canker disease. Full article

Thermostable cellulases and xylanases have broad acceptance in food, feed, paper and pulp, and bioconversion of lignocellulosics. Thermophilic fungi serve as an excellent source of thermostable enzymes. This study characterized four endo-β-1,4-glucanases (two glycoside hydrolase (GH) family 5 and two GH7 members) and four endo-β-1,4-xylanases (two GH10 and two GH11 members) from thermophilic fungus Thielavia terrestris, along with one GH10 endo-β-1,4-xylanase each from thermophilic fungus Chaetomium thermophilum and mesophilic fungus Chaetomium globosum. Comparative analysis was conducted against three previously reported GH10 endoxylanases: two thermostable enzymes from the thermophilic fungus Humicola insolens and thermophilic bacterium Halalkalibacterium halodurans, and one mesophilic enzyme from model fungus Neurospora crassa. The GH10 xylanase TtXyn10C (Thite_2118148; UniProt G2R8T7) from T. terrestris demonstrated high thermostability and activity, with an optimal temperature of 80–85 °C. It retained over 60% of its activity after 2 h at 70 °C, maintained approximately 30% activity after 15 min at 80 °C, and showed nearly complete stability following 1 min of exposure to 95 °C. TtXyn10C exhibited specific activity toward beechwood xylan (1130 ± 15 U/mg) that exceeded xylanases from H. insolens and H. halodurans while being comparable to N. crassa xylanase activity. Furthermore, TtXyn10C maintained stability across a pH range of 3–9 and resisted trypsin digestion, indicating its broad applicability. The study expands understanding of enzymes from thermophilic fungi. The discovery of the TtXyn10C offers a new model for investigating the high activity-stability trade-off and structure-activity relationships critical for industrial enzymes. Full article

The increasing emission of greenhouse gases that comes with the rise in industrialization is harmful to the environment. Thus, finding new renewable energy sources is becoming increasingly important in the energy field. One such renewable energy source is biomass, which provides valuable energy carriers—for example, biofuels. The objective of this work was to evaluate the release of total reducing sugars (TRSs) from mesquite pod seed hulls by chemical and enzymatic hydrolysis. The husks were crushed and separated by screens (#16, #30 and #50). The effect of hydrolysis time (10, 20, and 30 min) and sulfuric acid concentration (0, 0.25, and 0.5 N) was analyzed. The #50 mesh showed the highest TRS release, increasing from 3.19 to 17.49 g/L as the reaction time was extended. Additionally, enzymatic hydrolysis with endo-1, 4-β-xylanase and β-glucanase enzymes was evaluated on the solid and liquid fractions obtained. Statistical analysis with Design Expert showed that, for the solid fractions, after 31 h there were no significant differences, reaching 79.46 g/L TRS. In the liquid fractions, the TRS released reached 113.37 g/L after 54 h of enzymatic treatment. The release of TRS by chemical hydrolysis was also modeled with artificial neural networks, considering the particle size, the hydrolysis time, and the sulfuric acid concentration. The coefficient of determination (r²) indicates that the ANNs present a better data fit (r² > 0.99) to predict the experimental conditions that maximize the study variables. Full article

Background: Macrobrachium amazonicum is an opportunistic and omnivorous species that primarily feeds on plant material. Recent studies have shown that Endo-β-1,4-glucanase and Endo-β-1,4-mannanase are expressed in the transcriptome of adult specimens, while juveniles are capable of digesting nutrients from purified cellulose in their diet. In organisms that degrade raw plant material, laccase plays a key role in oxidizing phenolic compounds found in lignin, leading to its depolymerization and increasing access to cellulose and hemicellulose microfibrils. Objective: In this study, we conducted an in silico identification and characterization of the laccase-encoding gene, as this enzyme is linked to lignin biodegradation in herbivorous crustaceans. Methods: We analyzed the transcriptomes of the hepatopancreas from adult M. amazonicum, sequenced using the Illumina HiSeq 2500 platform. Subsequently, bioinformatics analyses were conducted to predict the conserved regions and active sites associated with laccase activity. Results: A complete open reading frame (ORF) of the laccase protein was identified in all datasets, comprising 609 amino acids. The top 40 similarity hits corresponded exclusively to crustaceans such as prawns, crayfish, and crabs (86.3–51.4%), while the highest divergence was observed in relation to fungi, plants, and bacteria. Three conserved domains were detected, along with the complete set of copper-binding centers (T1Cu, T2Cu, and T3Cu). A notable variable residue was methionine, suggesting a reduced redox potential in M. amazonicum laccase. Conclusion: These findings, combined with recent reports on the nutritional requirements of M. amazonicum, contribute to a deeper understanding of the digestive physiology of this species and offer valuable insights into its ability to utilize plant fibers as energy sources. Full article

To enhance the nutritional value of Acanthopanax senticosus leaves (AL), a fermentation process was conducted using a probiotic Bacillus mixture, and the changes in chemical constituents and biological activities before and after fermentation were compared. A response surface methodology was employed to optimize the liquid fermentation conditions of AL based on their influence on polyphenol content. Non-targeted metabolomics analysis was performed using LC-MS/MS to reveal the differing profiles of compounds before and after fermentation. The results indicated that Bacillus subtilis LK and Bacillus amyloliquefaciens M2 significantly influenced polyphenol content during fermentation. The optimal fermentation conditions were determined to be a fermentation time of 54 h, a temperature of 39.6 °C, and an inoculum size of 2.5% (v/v). In comparison to unfermented AL, the total polyphenol and flavonoid contents, as well as the free radical scavenging capacities measured by DPPH and ABTS assays, and the activities of β-glucosidase and endo-glucanase, were significantly increased. The non-targeted metabolomics analysis identified 1348 metabolites, of which 829 were classified as differential metabolites. A correlation analysis between the differential metabolites of polyphenols, flavonoids, and antioxidant activity revealed that 13 differential metabolites were positively correlated with antioxidant activity. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis of the differential metabolites identified 82 pathways, with two of the top 25 metabolic pathways related to flavonoids. This study explores the potential for enhancing the active ingredients and biological effects of AL through probiotic fermentation using Bacillus strains. Full article

This study explores the in vitro antifungal effects of nerol, a linear acyclic monoterpene alcohol of plant origin, on Fusarium oxysporum, Pestalotiopsis neglecta, and Valsa mali. To further investigate the antifungal mechanism of nerol against F. oxysporum, we examined changes in mycelial morphology and cell membrane integrity-related indices, as well as the activities of antioxidant and pathogenicity-related enzymes. The results demonstrated that nerol exhibited significant concentration-dependent inhibition of mycelial growth in all three fungi, with EC₅₀ values of 0.46 μL/mL for F. oxysporum, 1.81 μL/mL for P. neglecta, and 1.26 μL/mL for V. mali, with the strongest antifungal activity observed against F. oxysporum. Scanning electron microscopy revealed that nerol severely disrupted the mycelial structure of F. oxysporum, causing deformation, swelling, and even rupture. Treatment with 0.04 μL/mL nerol led to significant leakage of soluble proteins and intracellular ions in F. oxysporum, and the Na⁺/K⁺-ATPase activity was reduced to 28.02% of the control, indicating enhanced membrane permeability. The elevated levels of hydrogen peroxide and malondialdehyde, along with propidium iodide staining of treated microconidia, further confirmed cell membrane disruption caused by nerol. Additionally, after 12 h of exposure to 0.04 μL/mL nerol, the activity of superoxide dismutase in F. oxysporum decreased to 55.81% of the control, and the activities of catalase and peroxidase were also significantly inhibited. Nerol markedly reduced the activities of pathogenicity-related enzymes, such as endo-1,4-β-D-glucanase, polygalacturonase, and pectin lyase, affecting fungal growth and virulence. In conclusion, nerol disrupts the cell membrane integrity and permeability of F. oxysporum, reduces its virulence, and ultimately inhibits fungal growth, highlighting its potential as an alternative to chemical fungicides for controlling F. oxysporum. Full article

Enzyme-production microorganisms typically occupy a dominant position in composting, where cellulolytic microorganisms actively engage in the breakdown of lignocellulose. Exploring strains with high yields of cellulose-degrading enzymes holds substantial significance for the industrial production of related enzymes and the advancement of clean bioenergy. This study was inclined to screen cellulolytic bacteria, conduct genome analysis, mine cellulase-related genes, and optimize cellulase production. The potential carboxymethylcellulose-hydrolyzing bacterial strain Z2.6 was isolated from the maturation phase of pig manure-based compost with algae residuals as the feedstock and identified as Bacillus velezensis. In the draft genome of strain Z2.6, 31 related cellulolytic genes were annotated by the CAZy database, and further validation by cloning documented the existence of an endo-1,4-β-D-glucanase (EC 3.2.1.4) belonging to the GH5 family and a β-glucosidase (EC 3.2.1.21) belonging to the GH1 family, which are predominant types of cellulases. Through the exploration of ten factors in fermentation medium with Plackett–Burman and Box–Behnken design methodologies, maximum cellulase activity was predicted to reach 2.98 U/mL theoretically. The optimal conditions achieving this response were determined as 1.09% CMC-Na, 2.30% salinity, and 1.23% tryptone. Validation under these specified conditions yielded a cellulose activity of 3.02 U/mL, demonstrating a 3.43-fold degree of optimization. In conclusion, this comprehensive study underscored the significant capabilities of strain Z2.6 in lignocellulolytic saccharification and its potentialities for future in-depth exploration in biomass conversion. Full article

Meloidogyne javanica is one of the most widespread and economically important sedentary endoparasites. In this study, a comparative transcriptome analysis of M. javanica between pre-parasitic second-stage juveniles (Pre-J2) and parasitic juveniles (Par-J3/J4) was conducted. A total of 48,698 unigenes were obtained, of which 18,826 genes showed significant differences in expression (p < 0.05). In the differentially expressed genes (DEGs) from transcriptome data at Par-J3/J4 and Pre-J2, a large number of unigenes were annotated to the C-type lectin (CTL, Mg01965), the cathepsin L-like protease (Mi-cpl-1), the venom allergen-like protein (Mi-mps-1), Map-1 and the cellulase (endo-β-1,4-glucanase). Among seven types of lectins found in the DEGs, there were 10 CTLs. The regulatory roles of Mj-CTL-1, Mj-CTL-2 and Mj-CTL-3 in plant immune responses involved in the parasitism of M. javanica were investigated. The results revealed that Mj-CTL-2 could suppress programmed cell death (PCD) triggered by Gpa2/RBP-1 and inhibit the flg22-stimulated ROS burst. In situ hybridization and developmental expression analyses showed that Mj-CTL-2 was specifically expressed in the subventral gland of M. javanica, and its expression was up-regulated at Pre-J2 of the nematode. In addition, in planta silencing of Mj-CTL-2 substantially increased the plant resistance to M. javanica. Moreover, yeast co-transformation and bimolecular fluorescence complementation assay showed that Mj-CTL-2 specifically interacted with the Solanum lycopersicum catalase, SlCAT2. It was demonstrated that M. javanica could suppress the innate immunity of plants through the peroxide system, thereby promoting parasitism. Full article

Solid by-products with lignocellulosic structures are considered appropriate substrates for solid-state fermentation (SSF) to produce enzymes with diverse industrial applications. In this work, brewer’s spent grain (BSG), rice husk (RH), and vine shoot trimmings (VSTs) were employed as substrates in SSF with Aspergillus niger CECT 2088 to produce cellulases, xylanases, and amylases. The addition of 2% (NH₄)₂SO₄ and 1% K₂HPO₄ to by-products had a positive effect on enzyme production. Substrate particle size influenced enzyme activity and the overall highest activities were achieved at the largest particle size (10 mm) of BSG and RH and a size of 4 mm for VSTs. Optimal substrate composition was predicted using a simplex centroid mixture design. The highest activities were obtained using 100% BSG for β-glucosidase (363 U/g) and endo-1,4-β-glucanase (189 U/g), 87% BSG and 13% RH for xylanase (627 U/g), and 72% BSG and 28% RH for amylase (263 U/g). Besides the optimal values found, mixtures of BSG with RH or VSTs proved to be alternative substrates to BSG alone. These findings demonstrate that SSF bioprocessing of BSG individually or in mixtures with RH and VSTs is an efficient and sustainable strategy to produce enzymes of significant industrial interest within the circular economy guidelines. Full article

The purpose of this study was to study the chemical composition, rumen degradation characteristics, surface attached microbial community and cellulase activity of garlic skin (GS) and Artemisia argyi stalk (AS), in order to explain their feeding value. Four 14-month-old healthy Min Dong male goats with permanent rumen fistula were selected as experimental animals. The rumen degradation characteristics of GS and AS were determined by using the nylon bag method, and the bacterial composition, cellulase activity and their relationship on the surface of the two groups were analyzed with high-throughput sequencing of 16S rRNA gene. The results showed that in GS and AS, the effective degradation rate (ED) values of dry matter (DM) were 42.53% and 37.12%, the ED values of crude protein (CP) were 37.19% and 43.38%, the ED values of neutral detergent fiber (NDF) were 36.83% and 36.23%, and the ED values of acid detergent fiber (ADF) were 33.81% and 34.77%. During rumen degradation, the richness and evenness of bacteria attached to the AS surface were higher. At the phylum level, Bacteroidetes and Firmicutes were always the main rumen bacteria in the two groups. At the genus level, fiber-degrading bacteria such as Prevotella, Treponema, and Ruminococcus showed higher levels in GS (p < 0.05). Compared with GS, the activity of β-glucosidase (BG enzyme), endo-β-1,4-glucanase (C1 enzyme), exo-β-1,4-glucanase (Cx enzyme) and neutral xylanase (NEX enzyme) attached to AS surface showed a higher trend. Correlation analysis showed that the relative abundance of Succinivibrio and Rikenellaceae_RC9_gut_group was positively correlated with the rumen degradability of nutrients in GS, and the relative abundance of Christensenellaceae R-7_group, Succinivibrio and Ruminococcus was positively correlated with the rumen degradability of nutrients in AS. The conclusion of this study shows that AS has more potential to become ruminant roughage than GS. In addition, this study also revealed the relationship between cellulase activity and bacteria, which provided new information for us to better analyze the effects of GS and AS on the rumen of ruminants and provided an important theoretical basis for the development and utilization of agricultural by-products. Full article

Diosgenin is an important raw material used in the synthesis of steroid drugs, and it is widely used in the pharmaceutical industry. The traditional method of producing diosgenin is through using raw materials provided via the plant Dioscorea zingiberensis C. H. Wright (DZW), which is subsequently industrially hydrolyzed using a high quantity of hydrochloric and sulfuric acids at temperatures ranging from 70 °C to 175 °C. This process results in a significant amount of unmanageable wastewater, creates issues of severe environmental pollution and consumes high quantities of energy. As an alternative, the enzymolysis of DZW to produce diosgenin is an environmentally and friendly method with wide-ranging prospects for its application. However, there are still only a few enzymes that are suitable for production on an industrial scale. In this study, three new key enzymes, E1, E2, and E3, with a high conversion stability of diosgenin, were isolated and identified using an enzyme-linked-substrate autography strategy. HPLC-MS/MS identification showed that E1, a 134.45 kDa protein with 1019 amino acids (AAs), is a zinc-dependent protein similar to the M16 family. E2, a 97.89 kDa protein with 910 AAs, is a type of endo-β-1,3-glucanase. E3, a 51.6 kDa protein with 476 AAs, is a type of Xaa-Pro aminopeptidase. In addition, the method to immobilize these proteins was optimized, and stability was achieved. The results show that the optimal immobilization parameters are 3.5% sodium alginate, 3.45% calcium chloride concentration, 1.4 h fixed time, and pH 8.8; and the recovery rate of enzyme activity can reach 43.98%. A level of 70.3% relative enzyme activity can be obtained after employing six cycles of the optimized technology. Compared with free enzymes, immobilized enzymes have improved stability, acid and alkaline resistance and reusability, which are conducive to large-scale industrial production. Full article

β-1,3-glucanase plays an important role in the biodegradation, reconstruction, and development of β-1,3-glucan. An endo-β-1,3-glucanase which was encoded by PeBgl1 was expressed, purified and characterized from Penicillium expansum for the first time. The PeBgl1 gene was amplified and transformed into the competent cells of E. coli Rosetta strain with the help of the pET-30a cloning vector. The recombinant protein PeBgl1 was expressed successfully at the induction conditions of 0.8 mmol/L IPTG at 16 °C for 16 h and then was purified by nickel ion affinity chromatography. The optimum reaction temperature of PeBgl1 was 55 °C and it had maximal activity at pH 6.0 according to the enzymatic analysis. Na₂HPO₄-NaH₂PO₄ buffer (pH 6.0) and NaCl have inhibitory and enhancing effects on the enzyme activities, respectively. SDS, TritonX-100 and some metal ions (Mg²⁺, Ca²⁺, Ba²⁺, Cu²⁺, and Zn²⁺) have an inhibitory effect on the enzyme activity. The results showed that PeBgl1 protein has good enzyme activity at 50–60 °C and at pH 5.0–9.0, and it is not a metal dependent enzyme, which makes it robust for storage and transportation, ultimately holding great promise in green biotechnology and biorefining. Full article

In this study, we investigated the effect of exogenous melatonin (MT) on cell wall metabolism leading to Chinese plum (Prunus salicina Lindl.) fruit softening. Exogenous MT treatment increased the endogenous MT content in plum fruits before fruit ripening. However, in mature plum fruits, exogenous MT treatment decreased the fruit hardness, pulp hardness, fruit elasticity, contents of ion-bound pectin, covalently-bound pectin, hemicellulose, and cellulose, and activities of xyloglucan endotransglycosylase/hydrolase and endo-β-1,4-glucanase, and increased the water-soluble pectin content, and activities of pectin methyl esterase, pectin lyase, polygalacturonase, β-galactopyranosidase, and α-L-arabinofuranosidase. Transcriptome analysis revealed that the differentially expressed genes (DEGs) associated with cell wall metabolism in the exogenous MT-treated plum fruits were mainly enriched in the pentose and glucuronate interconversions, phenylpropanoid biosynthesis, cyanoamino acid metabolism, and galactose metabolism pathways. Analysis of these DEGs revealed that exogenous MT treatment affected the expression of genes regulating the cell wall metabolism. Overall, exogenous MT treatment promotes the fruit softening of Chinese plum. Full article

Xanthan is an extracellular heteropolysaccharide produced by the bacteria Xanthomonas campestris. Due to its unique properties, the polysaccharide and its derivatives are widely used in many industries, from food to biomedicine and oil production, that demands an efficient xanthan depolymerization method to adapt this polysaccharide for various applications. Unlike the known chemical approaches, biological methods are considered to be more environmentally friendly and less energy intensive. In laboratory conditions, we have isolated a bacterial community capable of reducing the xanthan viscosity. Identification of the individual isolates in the microbial community and their testing resulted in the consortium LE-C1, consisting of two microorganisms Paenibacillus phytohabitans KG5 and Cellulosimicrobium cellulans KG3. The specific activities of the overall xanthanase and auxiliary enzymes that may be involved in the xanthan depolymerization were as follows: xanthanase, 19.6 ± 0.6 U/g; β-glucosidase, 3.4 ± 0.1 U/g; α-mannosidase, 68.0 ± 2.0 U/g; β-mannosidase, 0.40 ± 0.01 U/g; endo-glucanase, 4.0 ± 0.1 U/g; and xanthan lyase, 2.20 ± 0.07 U/mg. In order to increase the efficiency of xanthan biodegradation, the LE-C1 whole cells were immobilized in a poly(vinyl alcohol) cryogel. The resulting regenerative biocatalyst was able to complete xanthan depolymerization within 40 cycles without loss of activity or degradation of the matrix. Full article

Marennine, a blue pigment produced by the blue diatom Haslea ostrearia, is known to have some biological activities. This pigment is responsible for the greening of oysters on the West Coast of France. Other new species of blue diatom, H. karadagensis, H. silbo sp. inedit., H. provincialis sp. inedit, and H. nusantara, also produce marennine-like pigments with similar biological activities. Aside from being a potential source of natural blue pigments, H. ostrearia-like diatoms present a commercial potential for the aquaculture, food, cosmetics, and health industries. Unfortunately, for a hundred years, the exact molecular structure of this bioactive compound has remained a mystery. A lot of hypotheses regarding the chemical structure of marennine have been proposed. The recent discovery of this structure revealed that it is a macromolecule, mainly carbohydrates, with a complex composition. In this study, some glycoside hydrolases were used to digest marennine, and the products were further analyzed using nuclear magnetic resonance (NMR) and mass spectroscopy (MS). The reducing sugar assay showed that marennine was hydrolyzed only by endo-1,3-β-glucanase. Further insight into the structure of marennine was provided by the spectrum of ¹H NMR, MS, a colorimetric assay, and a computational study, which suggest that the chemical structure of marennine contains 1,3-β-glucan. Full article