Optimization, gene cloning, expression, and molecular docking insights for enhanced cellulase enzyme production by Bacillus amyloliquefaciens strain elh1.

BACKGROUND: In this study, we isolated a cellulase-producing bacterium, Bacillus amyloliquefaciens strain elh, from rice peel. We employed two optimization methods to enhance the yield of cellulase. Firstly, we utilized a one-variable-at-a-time (OVAT) approach to evaluate the impact of individual physical and chemical parameters. Subsequently, we employed response surface methodology (RSM) to investigate the interactions among these factors. We heterologously expressed the cellulase encoding gene using a cloning vectorin E. coli DH5α. Moreover, we conducted in silico molecular docking analysis to analyze the interaction between cellulase and carboxymethyl cellulose as a substrate. RESULTS: The bacterial isolate eh1 exhibited an initial cellulase activity of 0.141 ± 0.077 U/ml when cultured in a specific medium, namely Basic Liquid Media (BLM), with rice peel as a substrate. This strain was identified as Bacillus amyloliquefaciens strain elh1 through 16S rRNA sequencing, assigned the accession number OR920278 in GenBank. The optimal incubation time was found to be 72 h of fermentation. Urea was identified as the most suitable nitrogen source, and dextrose as the optimal sugar, resulting in a production increase to 5.04 ± 0.120 U/ml. The peak activity of cellulase reached 14.04 ± 0.42 U/ml utilizing statistical optimization using Response Surface Methodology (RSM). This process comprised an initial screening utilizing the Plackett-Burman design and further refinement employing the BOX -Behnken Design. The gene responsible for cellulase production, egl, was effectively cloned and expressed in E. coli DH5α. The transformed cells exhibited a cellulase activity of 22.3 ± 0.24 U/ml. The egl gene sequence was deposited in GenBank with the accession number PP194445. In silico molecular docking revealed that the two hydroxyl groups of carboxymethyl cellulose bind to the residues of Glu169 inside the binding pocket of the CMCase. This interaction forms two hydrogen bonds, with an affinity score of -5.71. CONCLUSIONS: Optimization of cultural conditions significantly enhances the yield of cellulase enzyme when compared to unoptimized culturing conditions. Additionally, heterologous expression of egl gene showed that the recombinant form of the cellulase is active and that a valid expression system can contribute to a better yield of the enzyme.

Enhanced flexibility of high-yield bamboo pulp fibers via cellulase immobilization within guar gum/polyacrylamide/polydopamine interpenetrating network hydrogels.

Softness is a crucial criterion in assessing the comfort and usability of tissue paper. Flexible fibers contribute to the softness of the tissue paper by allowing the sheets to conform to the contours of the skin without feeling rough or abrasive. This study focuses on developing innovative CGG/APAM/PDA hydrogels with interpenetrating networks consisting of cationic guar gum, anionic polyacrylamide, and polydopamine for cellulase immobilization, aimed at improving bamboo fiber flexibility. Cellulase biomolecules are efficiently immobilized on CGG/APAM/PDA hydrogels through the Schiff base reaction. Immobilized cellulases have a wider pH applicability than free cellulases, good storage stability, and can maintain high relative activity at relatively high temperatures. The treatment of bamboo fibers with immobilized cellulase results in a significant increase in flexibility, reaching 6.90 × 1014 N·m2, which is 7.18 times higher than that of untreated fibers. The immobilization of cellulases using CGG/APAM/PDA hydrogels as carriers results in a substantial enhancement of storage stability, pH applicability, and inter-fiber bonding strength, as well as the capacity to sustain high relative enzymatic activity at elevated temperatures. The immobilization of cellulase within CGG/APAM/PDA interpenetrating network hydrogels presents a viable strategy for enhancing bamboo fiber flexibility, thereby expanding the accessibility of tissue products.

Mass production and characterization of an endoglucanase from Coleoptera insect (Monochamus saltuarius) in yeast Kluyveromyces lactis.

To harness the diverse industrial applications of cellulase, including its use in the food, pulp, textile, agriculture, and biofuel sectors, this study focused on the high-yield production of a bioactive insect-derived endoglucanase, Monochamus saltuarius GH Family 5 (MsGHF5). MsGHF5 was introduced into the genome of Kluyveromyces lactis to maintain expression stability, and mass production of the enzyme was induced using fed-batch fermentation. After 40 h of cultivation, recombinant MsGHF5 was successfully produced in the culture broth, with a yield of 29,000 U/L, upon galactose induction. The optimal conditions for the activity of purified MsGHF5 were determined to be a pH of 5 and a temperature of 35 °C, with the presence of ferrous ions enhancing the enzymatic activity by up to 1.5-fold. Notably, the activity of MsGHF5 produced in K. lactis was significantly higher than that produced in Escherichia coli, suggesting that glycosylation is crucial for the functional performance of the enzyme. This study highlights the potential use of K. lactis as a host for the production of bioactive MsGHF5, thus paving the way for its application in various industrial sectors.

Enhancing dark fermentative hydrogen production from wheat straw through synergistic effects of active electric fields and enzymatic hydrolysis pretreatment.

Hydrogen, a clean and sustainable energy source, faces challenges from energy-intensive pre-processing technologies. This study explores the synergistic enhancement of active electric fields on enzymolysis of wheat straw and hydrogen production through dark fermentation. The active electric field enzymolysis system improved the adsorption capacity of wheat straw to cellulase, increasing cellulase activity by 18.0 %, causing a 39.1 % increase in reducing sugar content. In the active fermentation system, Clostridium_sensu_stricto_1 activity was enhanced in the first stage, increasing hydrogenase activity by 23.0 %, prolonging the first hydrogen production peak. Elevated reducing sugars were observed in the second stage, with Prevotella_9 and Bacteroides becoming the dominant hydrogen-producing bacteria in the third stage, leading to a second hydrogen production peak. Overall, cumulative hydrogen production was enhanced by 50.9 % compared to the control. The synergistic pretreatment with an active electric field and cellulase provides a novel approach for efficiently utilizing wheat straw.

Screening, characterization, and optimization of the fermentation conditions of a novel cellulase-producing microorganism from soil of Qinghai-Tibet Plateau.

Cellulases play an important role in the bioconversion of lignocellulose. Microorganisms found in extreme environments are a potentially rich source of cellulases with unique properties. Due to the uniqueness of the environment, the abundant microbial resources in the Qinghai-Tibet Plateau (QTP) are worth being explored. The aim of this study was to isolate and characterize an acidic, mesophilic cellulase-producing microorganism from QTP. Moreover, the fermentation conditions for cellulase production were optimized for future application of cellulase in the development of lignocellulose biomass. A novel cellulase-producing strain, Penicillium oxalicum XC10, was isolated from the soil of QTP. The cellulase produced by XC10 was a mesophilic cellulase that exhibited good acid resistance and some cold-adaptation properties, with maximum activity at pH 4.0 and 40°C. Cellulase activity was significantly enhanced by Na+ (p < 0.05) and inhibited by Mg2+, Ca2+, Cu2+, and Fe3+ (p < 0.05). After optimization, maximum cellulase activity (8.56 U/mL) was increased nearly 10-fold. Optimal fermentation conditions included an inoculum size of 3% (v/v) in a mixture of corn straw (40 g/L), peptone (5 g/L), and Mg2+ (4 g/L), at pH 4.0, 33°C, and shaking at 200 rpm. The specific properties of the P. oxalicum XC10 cellulase suggest the enzyme may serve as an excellent candidate for the bioconversion and utilization of lignocellulose biomass generated as agricultural and food-processing wastes.

Evaluating the reinforcing potential of enzymatic cellulose nanocrystals in polypropylene nanocomposite.

Cellulose nanocrystals (CNCs) produced through enzymatic hydrolysis exhibit physicochemical properties that make them attractive as eco-friendly reinforcing agents in polymer composites. However, the extent of their efficacy within a polymeric matrix is yet to be fully established. This study investigated the reinforcing capabilities of enzymatic CNC (approximately 3 nm in diameter) isolated from bleached eucalyptus Kraft pulp (BEKP), focusing on its application in polypropylene (PP) nanocomposites produced by injection molding. The study compared the performance of this enzymatic CNC (1-5 % wt) with PP composites reinforced with micro-sized cellulose fibers (BEKP at 10-30 % wt, approximately 13 µm) and additionally with commercial CNC produced by sulfuric acid hydrolysis. Despite enzymatic CNC experiencing agglomeration during spray-drying, leading to an average diameter increase to 3 µm, it still significantly increased the crystallization and glass transition temperature of the PP matrix. However, this agglomeration likely hindered the improvement of the mechanical properties within the nanocomposites. The results also showed that enzymatic CNC provided higher thermal stability at lower reinforcement levels compared to BEKP, but this came with a reduction in stiffness, posing a significant consideration in composite design. The addition of a coupling agent greatly enhanced the dispersion of reinforcements and the interfacial adhesion within the matrix, contributing to the enhanced performance of the composite properties. Additionally, enzymatic CNC demonstrated potential for superior reinforcement efficacy compared to commercially available CNC produced by sulfuric acid hydrolysis. In conclusion, enzymatic CNC exhibited a promising role as nano-reinforcement for thermoplastic polymer nanocomposites, exhibiting higher thermal properties at lower reinforcing loads than traditional micro-sized fiber reinforcements. The absence of sulfur, coupled with its higher thermal stability and sustainable potential, positions enzymatic CNC as a particularly favorable choice for applications involving direct contact with food, cosmetics, pharmaceuticals, and biomedical materials.

Effect of cellulose reducing ends and primary hydroxyl groups modifications on cellulose-cellulase interactions and cellulose hydrolysis.

Cellulose reducing ends are believed to play a vital role in the cellulose recalcitrance to enzymatic conversion. However, their role in insoluble cellulose accessibility and hydrolysis is not clear. Thus, in this study, reducing ends of insoluble cellulose derived from various sources were modified by applying reducing and/or oxidizing agents. The effects of cellulose reducing ends modification on cellulose reducing ends, cellulose structure, and cellulose accessibility to cellulase were evaluated along with the impact on cellulose hydrolysis with complete as well purified cellulase components. Sodium borohydride (NaBH4) reduction and sodium chlorite-acetic acid (SC/AA) oxidation were able to modify more than 90% and 60% of the reducing ends, respectively, while the bicinchoninic acid (BCA) reagent applied for various cycles oxidized cellulose reducing ends to various extents. X-ray diffractograms of the treated solids showed that these treatments did not change the cellulose crystalline structure and the change in crystallinity index was insignificant. Surprisingly, it was found that the cellulose reducing ends modification, either through selective NaBH4 reduction or BCA oxidation, had a negligible impact on cellulose accessibility as well on cellulose hydrolysis rates or final conversions with complete cellulase at loadings as low as 0.5 mg protein/g cellulose. In fact, in contrast to what is traditionally believed, modifications of cellulose reducing ends by these two methods had no apparent impact on cellulose conversion with purified cellulase components and their synergy. However, SC/AA oxidation resulted in significant drop in cellulose conversion (10%-50%) with complete as well purified cellulase components. Nonetheless, further research revealed that the cause for drop in cellulose conversion for the SC/AA oxidation case was due to primary hydroxyl groups (PHGs) oxidation and not the oxidation of reducing ends. Furthermore, it was found that the PHGs modification affects cellulose accessibility and slows the cellulase uptake as well resulting in significant drop in cellulose conversions.

Amylase and Cellulase Production from Newly Isolated Bacillus subtilis Using Acid Treated Potato Peel Waste.

Species belonging to the genus Bacillus produce many advantageous extracellular enzymes that have tremendous applications on a commercial scale for the textile, detergent, feed, food, and beverage industries. This study aimed to isolate potent thermo-tolerant amylolytic and cellulolytic bacterium from the local environment. Using the Box-Behnken design of response surface methodology, we further optimized the amylase and cellulase activity. The isolate was identified by 16S rRNA gene sequencing as Bacillus subtilis QY4. This study utilized potato peel waste (PPW) as the biomaterial, which is excessively being dumped in an open environment. Nutritional status of the dried PPW was determined by proximate analysis. All experimental runs were carried out in 250 mL Erlenmeyer flasks containing acid treated PPW as a substrate by the thermos-tolerant Bacillus subtilis QY4 incubated at 37 °C for 72 h of submerged fermentation. Results revealed that the dilute H2SO4 assisted autoclaved treatment favored more amylase production (0.601 IU/mL/min) compared to the acid treatment whereas high cellulase production (1.269 IU/mL/min) was observed in the dilute acid treatment and was found to be very effective compared to the acid assisted autoclaved treatment. The p-value, F-value, and coefficient of determination proved the significance of the model. These results suggest that PPW could be sustainably used to produce enzymes, which offer tremendous applications in various industrial arrays, particularly in biofuel production.

Cellulase production by Aspergillus fumigatus A4112 and the potential use of the enzyme in cooperation with surfactant to enhance floating oil recovery and methane production from palm oil mill effluent.

This research performed cellulase production by Aspergillus fumigatus A4112 and evaluated its potential use in palm oil mill effluent (POME) hydrolysis to recover oil simultaneously with the generation of fermentable sugar useful for biofuel production under non-sterilized conditions. Empty fruit bunch (EFB) without pretreatment was used as carbon source. The combination of nitrogen sources facilitated CMCase production. The maximum activity (3.27 U/mL) was obtained by 1.0 g/L peptone and 1.5 g/L (NH4)2SO4 and 20 g/L EFB at 40 °C for 7 days. High level of FPase activity (39.51 U/mL) was also obtained. Interestingly, the enzyme retained its cellulase activities more than 60% at ambient temperature over 15 days. In enzymatic hydrolysis, Triton X-100 was an effective surfactant to increase total oil recovery in the floating form. High yield of reducing sugar (50.13 g/L) and 21% (v/v) of floating oil was recoverable at 65 °C for 48 h. Methane content of the raw POME increased from 41.49 to 64.94% by using de-oiled POME hydrolysate which was higher than using the POME hydrolysate (59.82%). The results demonstrate the feasibility of the constructed process for oil recovery coupled with a subsequent step for methane yield enhancement in biogas production process that benefits the palm oil industry.

Draft genome sequence data on Bacillus safensis U41 isolated from soils of Santiniketan, India.

The draft genome sequence of an isolate of Bacillus safensis U41 from the soils of Santiniketan (23040'12″ N and 87039'52″ E) is reported here. Bacillus safensis is a bacterium that produces cellulases, which is essential for the breakdown of plant biomass. As such, it is a valuable source of digestive enzymes from plant biomass, especially cellulases. The genomic DNA was extracted from a single colony using a QIAgen Blood and Tissue kit (QIAgen Inc., Canada). Sequencing was performed via Illumina HiSeq X using 2 × 150 paired-end chemistry, generating 7,352,576 reads with sequence coverage of 509x. The assembly produced 20 contigs over 200 base pairs (bp) in length, with an N50 value of 901304 and an L50 of 2. The genome size was 3,732,407 bp, and the average GC content was 41.43 %. Genome annotation and gene predictions were performed using Prokka v.1.14.6, which identified 3783 coding sequences, 64 tRNA genes, and 3 rRNA genes.

Exploring the cellulolytic activity of environmental mycobacteria.

Although studies on non-tuberculous mycobacteria have increased in recent years because they cause a considerable proportion of infections, their cellulolytic system is still poorly studied. This study presents a characterization of the cellulolytic activities of environmental mycobacterial isolates derived from soil and water samples from the central region of Argentina, aimed to evaluate the conservation of the mechanism for the degradation of cellulose in this group of bacteria. The molecular and genomic identification revealed identity with Mycolicibacterium septicum. The endoglucanase and total cellulase activities were assessed both qualitatively and quantitatively and the optimal enzymatic conditions were characterized. A specific protein of around 56 kDa with cellulolytic activity was detected in a zymogram. Protein sequences possibly arising from a cellulase were identified by mass spectrometry-based shotgun proteomics. Results showed that M. septicum encodes for cellulose- and hemicellulose-related degrading enzymes, including at least an active ß-1,4 endoglucanase enzyme that could be useful to improve its survival in the environment. Given the important health issues related to mycobacteria, the results of the present study may contribute to the knowledge of their cellulolytic system, which could be important for their ability to survive in many different types of environments.

Heteroxylan hydrolysis by a recombinant cellulase-free GH10 xylanase from the alkaliphilic bacterium Halalkalibacterium halodurans C-125.

The search for affordable enzymes with exceptional characteristics is fundamental to overcoming industrial and environmental constraints. In this study, a recombinant GH10 xylanase (Xyn10-HB) from the extremely alkaliphilic bacterium Halalkalibacterium halodurans C-125 cultivated at pH 10 was cloned and expressed in E. coli BL21(DE3). Removal of the signal peptide improved the expression, and an overall activity of 8 U/mL was obtained in the cell-free supernatant. The molecular weight of purified Xyn10-HB was estimated to be 42.6 kDa by SDS-PAGE. The enzyme was active across a wide pH range (5-10) with optimal activity recorded at pH 8.5 and 60 °C. It also presented good stability with a half-life of 3 h under these conditions. Substrate specificity studies showed that Xyn10-HB is a cellulase-free enzyme that conventionally hydrolyse birchwood and oat spelts xylans (Apparent Km of 0.46 mg/mL and 0.54 mg/mL, respectively). HPLC analysis showed that both xylans hydrolysis produced xylooligosaccharides (XOS) with a degree of polymerization (DP) ranging from 2 to 9. The conversion yield was 77% after 24 h with xylobiose and xylotriose as the main end-reaction products. When assayed on alkali-extracted wheat straw heteroxylan, the Xyn10-HB produced active XOS with antioxidant activity determined by the DPPH radical scavenging method (IC50 of 0.54 mg/mL after 4 h). Owing to its various characteristics, Xyn10-HB xylanase is a promising candidate for multiple biotechnological applications.

Enhancing cellulose hydrolysis via cellulase immobilization on zeolitic imidazolate frameworks using physical adsorption.

This study investigates the immobilization of cellulase on zeolitic imidazolate frameworks (ZIFs) by physical adsorption, specifically the ZIF-8-NH2 and Fe3O4@ZIF-8-NH2, to enhance enzymatic hydrolysis efficiency. The immobilization process was thoroughly analyzed, including optimization of conditions and characterization of ZIF carriers and immobilized enzymes. The impacts on the catalytic activity of cellulase under various temperatures, pH levels, and storage conditions were examined. Additionally, the reusability of the immobilized enzyme was assessed. Results showed the cellulase immobilized on Fe3O4@ZIF-8-NH2 exhibited a high loading capacity of 339.64 mg/g, surpassing previous studies. Its relative enzymatic activity was found to be 71.39%. Additionally, this immobilized enzyme system demonstrates robust reusability, retaining 68.42% of its initial activity even after 10 cycles. These findings underscore the potential of Fe3O4@ZIF-8-NH2 as a highly efficient platform for cellulase immobilization, with promising implications for lignocellulosic biorefinery.

Composition of Lignocellulose Hydrolysate in Different Biorefinery Strategies: Nutrients and Inhibitors.

The hydrolysis and biotransformation of lignocellulose, i.e., biorefinery, can provide human beings with biofuels, bio-based chemicals, and materials, and is an important technology to solve the fossil energy crisis and promote global sustainable development. Biorefinery involves steps such as pretreatment, saccharification, and fermentation, and researchers have developed a variety of biorefinery strategies to optimize the process and reduce process costs in recent years. Lignocellulosic hydrolysates are platforms that connect the saccharification process and downstream fermentation. The hydrolysate composition is closely related to biomass raw materials, the pretreatment process, and the choice of biorefining strategies, and provides not only nutrients but also possible inhibitors for downstream fermentation. In this review, we summarized the effects of each stage of lignocellulosic biorefinery on nutrients and possible inhibitors, analyzed the huge differences in nutrient retention and inhibitor generation among various biorefinery strategies, and emphasized that all steps in lignocellulose biorefinery need to be considered comprehensively to achieve maximum nutrient retention and optimal control of inhibitors at low cost, to provide a reference for the development of biomass energy and chemicals.

Introduction of Cellulolytic Bacterium Bacillus velezensis Z2.6 and Its Cellulase Production Optimization.

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-ß-D-glucanase (EC 3.2.1.4) belonging to the GH5 family and a ß-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.

Platycodon grandiflorum saponins: Ionic liquid-ultrasound-assisted extraction, antioxidant, whitening, and antiaging activity.

This study explored the use of ionic liquid-ultrasound (ILU)-assisted extraction to enhance the extraction rate of Platycodon grandiflorum saponins (PGSs), and the content, extraction mechanism, antioxidant activity, whitening, and antiaging activity of PGSs prepared using ILU, ultrasound-water, thermal reflux-ethanol, and cellulase hydrolysis were compared. The ILU method particularly disrupted the cell wall, improved PGS extraction efficiency, and yielded a high total saponin content of 1.45 ± 0.02 mg/g. Five monomeric saponins were identified, with platycodin D being the most abundant at 1.357 mg/g. PGSs displayed excellent in vitro antioxidant activity and exhibited inhibitory effects on tyrosinase, elastase, and hyaluronidase. The results suggest that PGSs may have broad antioxidant, skin-whitening, and antiaging potential to a large extent. Overall, this study provided valuable insights into the extraction, identification, and bioactivities of PGSs, which could serve as a reference for future development and application of these compounds in the functional foods industry.

Synthesis of Extracellular L-lysine-α-oxidase along with Degrading Enzymes by Trichoderma cf. aureoviride Rifai VKM F-4268D: Role in Biocontrol and Systemic Plant Resistance.

When cultivating on wheat bran or deactivated fungal mycelium as a model of "natural growth", the ability of Trichoderma to synthesize extracellular L-lysine-α-oxidase (LysO) simultaneously with cell-wall-degrading enzymes (proteases, xylanase, glucanases, chitinases, etc.), responsible for mycoparasitism, was shown. LysO, in turn, causes the formation of H2O2 and pipecolic acid. These compounds are known to be signaling molecules and play an important role in the induction and development of systemic acquired resistance in plants. Antagonistic effects of LysO have been demonstrated against phytopathogenic fungi and Gram-positive or Gram-negative bacteria with dose-dependent cell death. The antimicrobial effect of LysO decreased in the presence of catalase. The generating intracellular ROS in the presence of LysO was also shown in both bacteria and fungi, which led to a decrease in viable cells. These results suggest that the antimicrobial activity of LysO is due to two factors: the formation of exogenous hydrogen peroxide as a product of the enzymatic oxidative deamination of L-lysine and the direct interaction of LysO with the cell wall of the micro-organisms. Thus, LysO on its own enhances the potential of the producer in the environment; namely, the enzyme complements the strategy of the fungus in biocontrol and indirectly participates in inducing SAR and regulating the relationship between pathogens and plants.

Once upon a Time, There Was a Piece of Wood: Present Knowledge and Future Perspectives in Fungal Deterioration of Wooden Cultural Heritage in Terrestrial Ecosystems and Diagnostic Tools.

Wooden Cultural Heritage (WCH) represents a significant portion of the world's historical and artistic heritage, consisting of immovable and movable artefacts. Despite the expertise developed since ancient times to enhance its durability, wooden artefacts are inevitably prone to degradation. Fungi play a pivotal role in the deterioration of WCH in terrestrial ecosystems, accelerating its decay and leading to alterations in color and strength. Reviewing the literature of the last 25 years, we aimed to provide a comprehensive overview of fungal diversity affecting WCH, the biochemical processes involved in wood decay, and the diagnostic tools available for fungal identification and damage evaluation. Climatic conditions influence the occurrence of fungal species in threatened WCH, characterized by a prevalence of wood-rot fungi (e.g., Serpula lacrymans, Coniophora puteana) in architectural heritage in temperate and continental climates and Ascomycota in indoor and harsh environments. More efforts are needed to address the knowledge fragmentation concerning biodiversity, the biology of the fungi involved, and succession in the degradative process, which is frequently centered solely on the main actors. Multidisciplinary collaboration among engineers, restorers, and life sciences scientists is vital for tackling the challenges posed by climate change with increased awareness. Traditional microbiology and culture collections are fundamental in laying solid foundations for a more comprehensive interpretation of big data.

Trophic Position of the White Worm (Enchytraeus albidus) in the Context of Digestive Enzyme Genes Revealed by Transcriptomics Analysis.

To assess the impact of Enchytraeidae (potworms) on the functioning of the decomposer system, knowledge of the feeding preferences of enchytraeid species is required. Different food preferences can be explained by variations in enzymatic activities among different enchytraeid species, as there are no significant differences in the morphology or anatomy of their alimentary tracts. However, it is crucial to distinguish between the contribution of microbial enzymes and the animal's digestive capacity. Here, we computationally analyzed the endogenous digestive enzyme genes in Enchytraeus albidus. The analysis was based on RNA-Seq of COI-monohaplotype culture (PL-A strain) specimens, utilizing transcriptome profiling to determine the trophic position of the species. We also corroborated the results obtained using transcriptomics data from genetically heterogeneous freeze-tolerant strains. Our results revealed that E. albidus expresses a wide range of glycosidases, including GH9 cellulases and a specific digestive SH3b-domain-containing i-type lysozyme, previously described in the earthworm Eisenia andrei. Therefore, E. albidus combines traits of both primary decomposers (primary saprophytophages) and secondary decomposers (sapro-microphytophages/microbivores) and can be defined as an intermediate decomposer. Based on assemblies of publicly available RNA-Seq reads, we found close homologs for these cellulases and i-type lysozymes in various clitellate taxa, including Crassiclitellata and Enchytraeidae.

Ionic liquid-assisted pretreatment of lignocellulosic biomass using purified Streptomyces MS2A cellulase for bioethanol production.

In recent years, the process of producing bioethanol from lignocellulosic biomass through biorefining has become increasingly important. However, to obtain a high yield of ethanol, the complex structures in the feedstock must be broken down into simple sugars. A cost-effective and innovative method for achieving this is ionic liquid pre-treatment, which is widely used to efficiently hydrolyze the lignocellulosic material. The study aims to produce a significant profusion of bioethanol via catalytic hydrolysis of ionic liquid-treated lignocellulose biomass. The current study reports the purification of Streptomyces sp. MS2A cellulase via ultrafiltration and gel permeation chromatography. The kinetic parameters and the biochemical nature of the purified cellulase were analyzed for the effective breakdown of the EMIM[OAC] treated lignocellulose chain. The two-step cellulase purification resulted in 6.28 and 12.44 purification folds. The purified cellulase shows a Km value of 0.82 ± 0.21 mM, and a Vmax value of 85.59 ± 8.87 µmol min-1 mg-1 with the catalytic efficiency of 1.027 S-1. The thermodynamic parameters like ΔH, ΔS, and ΔG of the system were studied along with the thermal deactivation kinetics of cellulase. The optimal temperature and pH of the purified cellulase enzyme for hydrolysis was found to be 40 °C and 7. The rice husk and wheat husk used in this study were pretreated with the EMIM [OAC] ionic liquid and the change in the structure of lignocellulosic biomass was observed via HRSEM. The ionic liquid treated biomass showed the highest catalytic hydrolysis yield of 106.66 ± 0.19 mol/ml on the third day. The obtained glucose was fermented with Saccharomyces cerevisiae to yield 23.43 g of ethanol/l of glucose from the rice husk (RH) and 24.28 g of ethanol/l of glucose from the wheat husk (WH).