Trichoderma Reesei Research Articles

Biochar derived from straw residue prepared via combined pre-treatment designed for efficient removal of tetracycline hydrochloride and sulfadiazine sodium salt

In this study, straw residue (SR) was prepared from corn straw by a combined pre-treatment method that involved both microbial treatment (Myrothecium verrucaria, Aspergillus niger, and Trichoderma reesei) and treatment with ρ-toluenesulfonic acid. After pre-treatment, the cellulose content of the residues reached 79.3 %, 72.1 %, 83.5 %, and 85.2 %, respectively. The results indicated that Aspergillus niger and Trichoderma reesei effectively destroyed the corn stover structure, improving the efficiency of the subsequent treatment. Following carbonation and activation processes, the SRs were converted into a series of biochars (ACCC, ACMC, ACTC, and ACNC) with large specific surface areas (2343, 2219, 2693, 2672 m2 g−1). The prepared biochars demonstrated excellent performance in adsorption tests performed using tetracycline hydrochloride (TC) and sulfadiazine sodium salt (SDZ) as adsorption models. The maximum adsorption capacities recorded for TC (908, 1117, 1216, and 1189 mg/g) and SDZ (930, 965, 1033, and 1083 mg/g) were higher than most of the other adsorbents. Furthermore, the potential adsorption mechanisms included pore filling, π-π interactions, hydrogen bonding, and electrostatic attraction. Even after 5 test cycles, the biochar retained over 75 % of its adsorption performance, highlighting its strong potential for applications in removing antibiotics from water.

Enhanced nutritional profile of dehulled peas through fungal fermentation: Impacts on protein, starch, saponins, and total phenolic content

AbstractDehulled peas (DHP) are increasingly popular in food applications, but their integration in food products is limited due to high starch and fiber causing gelling and aggregation during cooking. Additionally, DHP contains saponins (secondary metabolite), contributing to bitter flavors, yet they also hold health benefits. We hypothesize that fungal fermentation could enhance DHP nutritional profile and integration into food products. This study evaluated the effects of six fungal organisms (Aspergillus niger [An], Aspergillus oryzae [Ao], Aureobasidium pullulans [Ap], Neurospora crassa [Nc], Rhizopus microspores var. oligosporus [Ro], Trichoderma reesei [Tr]) on DHP over 120 h of submerged fermentation, evaluating total phenolics, starch, saponins, crude proteins, and overall mass balance. Results from the study demonstrated notable changes post‐fermentation, including increased overall protein content and solubility, decreased starch content, reduced overall mass recovery, and elevated levels of total phenolics and saponins. Filamentous fungi exhibited a significant reduction in starch content, contributing to a substantial reduction in mass recovery (31%–60%) compared to the control. Unexpectedly, saponin concentrations increased (1.5 to 3 folds) during fermentation, possibly attributed to the breakdown of the substrate matrix and release of bound saponins. Total phenolic levels varied among microorganisms, with An and Nc demonstrating the highest increases (6 to 10 folds) as compared to the control. Overall, these findings point to fungal fermentation as a tool for adding value to yellow peas and other crops facing similar processing challenges. Further research is warranted to understand the health impacts and value of these enhancements.

A preliminary study on the degradation of AFB1 by Tenebrio molitor, Rhizopus oryzae and Trichoderma reesei

Abstract Recently, genus Aspergillus, a fungus capable of producing aflatoxins, secondary highly toxic metabolites, has spread to new areas. These areas became suitable habitats due to the recent climate changes. The use of aflatoxin-contaminated crops is a cause of great concern in guaranteeing food safety and is responsible for major economic losses along the supply chain. For this reason, several strategies have been investigated to utilize these contaminated products as a possible food or feed resource by reducing or eliminating their aflatoxin content, but with limited relevant success. The presented study was aimed to evaluate a combination of biological processes to use aflatoxin B1 contaminated crops for their reintroduction into the production chain. The high tolerance to AFB1 and the apparent lack of accumulation in yellow mealworm larvae, reared on wheat bran substrates, spiked with increasing AFB1 concentration (0, 125, 250, 500 μg/kg) to obtain proteins of high biological value. Subsequently, the aflatoxin-degrading capacity of Rhizopus oryzae and Trichoderma reesei was applied to insect breeding waste (frass) in a fermentation process to ensure further utilization of biohazardous frass as soil conditioner. Individually, each process proven to be able to reduce the AFB1 present by about 80%, while the combination of the two approaches ensured the total degradation of aflatoxin B1-contaminated substrate and frass, which resulted in the possible production of biomass, that could be used for the feed and agricultural industry.

Performance assessment and characterization of the biosorption of the azo dye reactive Red 2 on thermally inactivated biomass of Trichoderma reesei

The present study evaluated and characterized the performance in the biosorption of the dye azo Reactive Red 2 (RR2) on thermally sterilized biomass of the fungus Trichoderma reesei. An FTIR analysis of unloaded and loaded biomass with RR2 confirmed the interactions between dye molecules and functional groups on the fungal biomass surface. The decrease in the initial pH of 9 to 3 with an increase in the temperature of 20 to 40 °C improved the biosorption, reaching for RR2 a higher removal efficiency of 76.88% at pH 3 and 40 °C. A pseudo-second-order model explained the biosorption kinetics of RR2 (R2≥0.9869), and the intraparticle diffusion model showed that the biosorption process was initially controlled by diffusion rate and later by biomass surface saturation with the dye molecules. The Langmuir isotherm explained the biosorption equilibrium well (R2 ≥ 0.9241), achieving the maximum biomass biosorption capacity (qmax ) value at 61.72 mg g−1. The dimensionless separation factor values (0.0632 ≤ RL ≤ 0.0973) and the thermodynamic evaluation of the process (ΔG° ≤ −24.09 kJ mol−1, ΔH° ≥ 12.39 kJ mol−1, ΔS° ≥ 0.1265 kJ mol−1 K−1) demonstrate that the biosorption was favorable, reversible, spontaneous, and endothermic. The findings in the present study allow it to consider the biomass of Trichoderma reesei thermally sterilized as a biosorbent biomaterial that can be used for bioprocess design in removing the azo reactive dyes from textile wastewater.

Potato and dairy industry side streams as feedstock for fungal and plant cell cultures

Utilization of industrial side streams as nutrient source for cellular agriculture is a promising option to improve the sustainability of the production processes. The aim of the study was to evaluate usability of two liquid food industry side streams, potato cell fluid and acid whey, as nutrient source for production of food ingredients with plant cell cultures (arctic bramble (Rubus arcticus L.) and tobacco BY-2 (Nicotiana tabacum L.)) and filamentous fungi (Paecilomyces variotii, Rhizopus oligosporus and Trichoderma reesei). The side streams were found to contain various sugars and other potential nutrients suitable for the studied organisms. The side streams were used as a sole nutrient source (fungi) or as a replacement of selected nutrients in the growth media (fungi and plant cells) in flask scale cultivations. Acid whey was successfully used as a growth medium for filamentous fungi, but it inhibited plant cell growth presumably due to high organic acid content (14 g/l). Potato side stream was found suitable as a fungal and plant cell growth media supplement, and it was used together with glucose for filamentous fungi, or as a partial replacer of macronutrients for plant cells, in bioreactor cultivations yielding high fungal (up to 36 g/l dry weight) and good plant cell (9.5 g/l dry weight) biomass production. Analyses of the produced biomasses revealed good nutritional value in terms of amino acid (17–27 % of dry matter) and dietary fiber (23–30 % of dry matter) contents, giving good premises for utilization in human nutrition.

Feed additives supplementation: a potential strategy to ameliorate heat stress in sheep

Abstract Given a significant climate-flexible and socio-economic role in developing nations, environmental heat stress imposes a major financial impact on sheep production systems globally endangering their production, reproduction, and growth. In this regard, the adverse effects of heat stress on sheep production systems have to be addressed through adoption of effective heat alleviation measures like animal management, nutritional management and genetic interventions of which the nutritional interventions seems to be the most cost effective way to alleviate heat stress. Nutritional manipulation for heat stress alleviation in sheep involves the use of antioxidant supplements (Vitamin B; Vitamin E and Selenium; Selenium; Zinc sulphate and folic acid; Vitamin C, Vitamin E, Selenium and Zinc; Naringin; Opuntia ficus-indica f. inermis; Açai oil and Brown seaweed like Ascophyllum nodosum and Sargassum latifolium). Further, electrolyte supplements (Dietary Electrolyte Balance (DEB); Sodium bicarbonate and potassium bicarbonate; Sodium hydroxide) have a beneficial effect on thermal responses, respiratory activities, gas exchange parameters, rumen fermentation, blood buffering capacity and acid-base balance. The mineral mixture supplements (Mineral blocks; Mineral mixture and antioxidants; Chromium; Zinc) play a crucial role in increasing the efficiency of antioxidant defence system, immunity-related parameters, production, reproduction, feed digestibility and insulin sensitivity. Probiotic supplements (Lactobacillus acidophilus, Saccharomyces cervisiae, Propionibacterium freudenreichii, Lactobacillus casei, Enterococcus faecium, Lactobacillus lactis, Bacillus subtilis, Propionibacterium freudenreichii, Pediococcus cerevisiae, Megaspha eraelsdenii, Bacillus licheniformis, Aspergillus oryzae, Schizochytrium limacinum, Trichoderma reesei and Saccharomyces cerevisiae) improve lactational performance, dietary energy utilization and productivity. The probiotics (live Saccharomyces cerevisiae) and prebiotics (mannan oligosaccharide plus b-glucans) used in heat stress alleviation improve dietary energy utilisation. Furthermore, the vital role of herbal supplements (Rosemary, Cinnamon, Turmeric, Clove, Naringin, Chestnut tannins, Giloy stem powder, Curcumin, Rocket oil (watercress oil), Flaxseed, Cornus, Oregano, Thyme, Chamomile flowers, Moringa oleifera, Betaine) has been highlighted to promote feed intake, antioxidant status, growth performance, feed utilization, reproductive performance and immune response. Effective adoption of nutritional strategies can thus ensure sustainable sheep production in this changing climate scenario.

Characterization of a GDS(L)-like hydrolase from Pleurotus sapidus with an unusual SGNH motif

The GDS(L)-like lipase from the Basidiomycota Pleurotus sapidus (PSA_Lip) was heterologously expressed using Trichoderma reesei with an activity of 350 U L−1. The isoelectric point of 5.0 was determined by isoelectric focusing. The novel PSA_Lip showed only 23.8–25.1%, 25.5%, 26.6% and 28.4% identity to the previously characterized GDSL-like enzymes phospholipase, plant lipase, acetylcholinesterase and acetylxylan esterase, from the carbohydrate esterase family 16, respectively. Therefore, the enzyme was purified from the culture supernatant and the catalytic properties and the substrate specificity of the enzyme were investigated using different assays to reveal its potential function. While no phospholipase, acetylcholinesterase and acetylxylan esterase activities were detected, studies on the hydrolysis of ferulic acid methyl ester (~ 8.3%) and feruloylated carbohydrate 5-O-transferuloyl-arabino-furanose (~ 0.8%) showed low conversions of these substrates. By investigating the hydrolytic activity towards p-nitrophenyl-(pNP)-esters with various chain-lengths, the highest activity was determined for medium chain-length pNP-octanoate at 65 °C and a pH value of 8, while almost no activity was detected for pNP-hexanoate. The enzyme is highly stable when stored at pH 10 and 4 °C for at least 7 days. Moreover, using consensus sequence analysis and homology modeling, we could demonstrate that the PSA_Lip does not contain the usual SGNH residues in the actives site, which are usually present in GDS(L)-like enzymes.

On the pH Dependency of the Catalysis by a Lytic Polysaccharide Monooxygenase from the Fungus Trichoderma reesei

On the pH Dependency of the Catalysis by a Lytic Polysaccharide Monooxygenase from the Fungus <i>Trichoderma reesei</i>

Transgressive phenotypes from outbreeding between the Trichoderma reesei hyper producer RutC30 and a natural isolate.

The filamentous fungus Trichoderma reesei produces cellulolytic enzymes that are essential for the hydrolysis of lignocellulosic biomass into monomerics sugars. The filamentous fungus T. reesei produces cellulolytic enzymes that are essential for the hydrolysis of lignocellulosic biomass into monomerics sugars, which can in turn be fermented to produce second-generation biofuels and bioproducts. Production performance improvement, which is essential to reduce production cost, relies on classical mutagenesis and genetic engineering techniques. Although sexual reproduction is a powerful tool for improving domesticated species, it is often difficult to apply to industrial fungi since most of them are considered asexual. In this study, we demonstrated that outbreeding is an efficient strategy to optimize T. reesei. Crossing between a natural isolate and a mutagenized strain generated a biodiverse progeny with some offspring displaying transgressive phenotype for cellulase activities.

New Proline-Specific Endoprotease for Improved Colloidal Beer Stability and Gluten Reduction

Chill-haze is a well-known concern for brewers and most commonly involves polyphenols that interact with proline-rich hordein proteins in beers. An effective haze-suppressing effect has previously been established by a proline-specific endoprotease (PEP). In the present study, the protease action of a new Trichoderma reesei expressed PEP enzyme, available as the product BCLEAR™ was studied by assessing its impact on colloidal as well as foam stability and gluten reduction in beer. All malt beers treated with the BCLEAR™ enzyme or beers produced using BCLEAR™ at low dosage levels during fermentation, showed improved colloidal and foam stability when compared to the benchmark PEP investigated. In addition, gluten reduction was investigated using the BCLEAR™ enzyme and was found to have a very good correlation with the observed haze reduction, suggesting that the PEP enzyme dosage can accurately be determined from the calculated gluten content of the given raw materials. Thus, for optimal PEP enzyme use, the brew processes may most efficiently be controlled by following the gluten content, which was demonstrated with use of Lateral Flow Device (LFD) analysis technology.

Upgraded cellulose and xylan digestions for synergistic enhancements of biomass enzymatic saccharification and bioethanol conversion using engineered Trichoderma reesei strains overproducing mushroom LeGH7 enzyme

Crop straws provide enormous lignocellulose resources transformable for sustainable biofuels and valuable bioproducts. However, lignocellulose recalcitrance basically restricts essential biomass enzymatic saccharification at large scale. In this study, the mushroom-derived cellobiohydrolase (LeGH7) was introduced into Trichoderma reesei (Rut-C30) to generate two desirable strains, namely GH7–5 and GH7–6. Compared to the Rut-C30 strain, both engineered strains exhibited significantly enhanced enzymatic activities, with β-glucosidases, endocellulases, cellobiohydrolases, and xylanase activities increasing by 113 %, 140 %, 241 %, and 196 %, respectively. By performing steam explosion and mild alkali pretreatments with mature straws of five bioenergy crops, diverse lignocellulose substrates were effectively digested by the crude enzymes secreted from the engineered strains, leading to the high-yield hexoses released for bioethanol production. Notably, the LeGH7 enzyme purified from engineered strain enabled to act as multiple cellulases and xylanase at higher activities, interpreting how synergistic enhancement of enzymatic saccharification was achieved for distinct lignocellulose substrates in major bioenergy crops. Therefore, this study has identified a novel enzyme that is active for simultaneous hydrolyses of cellulose and xylan, providing an applicable strategy for high biomass enzymatic saccharification and bioethanol conversion in bioenergy crops.

Fungal endo and exochitinase production, characterization, and application for Candida biofilm removal.

Chitinases are promising enzymes for a multitude of applications, including chitooligosaccharide (COS) synthesis for food and pharmaceutical uses and marine waste management. Owing to fungal diversity, fungal chitinases may offer alternatives for chitin degradation and industrial applications. The rapid reproduction cycle, inexpensive growth media, and ease of handling of fungi may also contribute to reducing enzyme production costs. Thus, this study aimed to identify fungal species with chitinolytic potential and optimize chitinase production by submerged culture and enzyme characterization using shrimp chitin. Three fungal species, Coriolopsis byrsina, Trichoderma reesei, and Trichoderma harzianum, were selected for chitinase production. The highest endochitinase production was achieved in C. byrsina after 168h cultivation (0.3 U mL- 1). The optimal temperature for enzyme activity was similar for the three fungal species (up to 45 and 55 ºC for endochitinases and exochitinases, respectively). The effect of pH on activity indicated maximum hydrolysis in acidic pH (4-7). In addition, the crude T. reesei extract showed promising properties for removing Candida albicans biofilms. This study showed the possibility of using shrimp chitin to induce chitinase production and enzymes that can be applied in different industrial sectors.

Safety evaluation of the food enzyme lysophospholipase from the genetically modified Trichoderma reesei strain DP-Nyc81.

The food enzyme lysophospholipase (2-lysophosphatidylcholine acylhydrolase, EC 3.1.1.5) is produced with the genetically modified Trichoderma reesei strain DP-Nyc81 by Genencor International B.V. The genetic modifications do not give rise to safety concerns. The food enzyme is free from viable cells of the production organism and its DNA. It is intended to be used in the processing of cereals and other grains for the production of glucose syrups and other starch hydrolysates. Since residual amounts of food enzyme-total organic solids are removed during these food manufacturing processes, dietary exposure was not calculated and toxicological studies were considered unnecessary. A search for the identity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that the risk of allergic reactions upon dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns, under the intended conditions of use.

Exploration of Trichoderma reesei as an alternative host for erythritol production

BackgroundErythritol, a natural polyol, is a low-calorie sweetener synthesized by a number of microorganisms, such as Moniliella pollinis. Yet, a widespread use of erythritol is limited by high production costs due to the need for cultivation on glucose-rich substrates. This study explores the potential of using Trichoderma reesei as an alternative host for erythritol production, as this saprotrophic fungus can be cultivated on lignocellulosic biomass residues. The objective of this study was to evaluate whether such an alternative host would lead to a more sustainable and economically viable production of erythritol by identifying suitable carbon sources for erythritol biosynthesis, the main parameters influencing erythritol biosynthesis and evaluating the feasibility of scaling up the defined process.ResultsOur investigation revealed that T. reesei can synthesize erythritol from glucose but not from other carbon sources like xylose and lactose. T. reesei is able to consume erythritol, but it does not in the presence of glucose. Among nitrogen sources, urea and yeast extract were more effective than ammonium and nitrate. A significant impact on erythritol synthesis was observed with variations in pH and temperature. Despite successful shake flask experiments, the transition to bioreactors faced challenges, indicating a need for further scale-up optimization.ConclusionsWhile T. reesei shows potential for erythritol production, reaching a maximum concentration of 1 g/L over an extended period, its productivity could be improved by optimizing the parameters that affect erythritol production. In any case, this research contributes valuable insights into the polyol metabolism of T. reesei, offering potential implications for future research on glycerol or mannitol production. Moreover, it suggests a potential metabolic association between erythritol production and glycolysis over the pentose phosphate pathway.

Enzymatic Production of Glycerol from Glycolaldehyde and Formaldehyde

Glycerol is a simple and important polyol in both biological and industrial field. To produce glycerol in an eco‐friendly manner, we established a one‐pot cascade reaction by fructose‐6‐phosphate aldolase (FSA) and aldehyde reductase (ALR) from formaldehyde (FALD) and its dimerized form glycolaldehyde (GALD) via L‐glyceraldehyde. For this, we characterized two FSAs at various conditions. Among them, FSA from Gilliamella apicola (GaFSA) has shown the higher L‐glyceraldehyde production activity in the phosphate buffer than the well‐known FSA from Escherichia coli (EcFSA) by more than 2 times. We also characterized two ALRs (xylose reductase from Candida tenuis; CtXR, the annotated DL‐glyceraldehyde reductase from Hypocrea jecorina; HjGLD) that specifically reduce L‐glyceraldehyde into glycerol. Bioproduction of glycerol from FALD and GALD was performed using GaFSA and CtXR, which produced ~8.8 mM glycerol from 25 mM of FALD and GALD (conversion yield of ~35%). This suggested that a simple process would help in designing a green production of glycerol from a wasteful one‐carbon chemicals.

Small GTPase Rab7 is involved in stress adaptation to carbon starvation to ensure the induced cellulase biosynthesis in Trichoderma reesei.

The saprophytic filamentous fungus Trichoderma reesei represents one of the most prolific cellulase producers. The bulk production of lignocellulolytic enzymes by T. reesei not only relies on the efficient transcription of cellulase genes but also their efficient secretion after being translated. However, little attention has been paid to the functional roles of the involved secretory pathway in the high-level production of cellulases in T. reesei. Rab GTPases are key regulators in coordinating various vesicle trafficking associated with the eukaryotic secretory pathway. Specifically, Rab7 is a representative GTPase regulating the transition of the early endosome to the late endosome followed by its fusion to the vacuole as well as homotypic vacuole fusion. Although crosstalk between the endosomal/vacuolar pathway and the secretion pathway has been reported, the functional role of Rab7 in cellulase production in T. reesei remains unknown. A TrRab7 was identified and characterized in T. reesei. TrRab7 was shown to play important roles in T. reesei vegetative growth and vacuole morphology. Whereas knock-down of Trrab7 significantly compromised the induced production of T. reesei cellulases, overexpression of the key transcriptional activator, Xyr1, restored the production of cellulases in the Trrab7 knock-down strain (Ptcu-rab7KD) on glucose, indicating that the observed defective cellulase biosynthesis results from the compromised cellulase gene transcription. Down-regulation of Trrab7 was also found to make T. reesei more sensitive to various stresses including carbon starvation. Interestingly, overexpression of Snf1, a serine/threonine protein kinase known as an energetic sensor, partially restored the cellulase production of Ptcu-rab7KD on Avicel, implicating that TrRab7 is involved in an energetic adaptation to carbon starvation which contributes to the successful cellulase gene expression when T. reesei is transferred from glucose to cellulose. TrRab7 was shown to play important roles in T. reesei development and a stress response to carbon starvation resulting from nutrient shift. This adaptation may allow T. reesei to successfully initiate the inducing process leading to efficient cellulase production. The present study provides useful insights into the functional involvement of the endosomal/vacuolar pathway in T. reesei development and hydrolytic enzyme production.

Optimizing microbioreactor cultivation strategies for Trichoderma reesei: from batch to fed-batch operations

BackgroundFilamentous fungi have long been recognized for their exceptional enzyme production capabilities. Among these, Trichoderma reesei has emerged as a key producer of various industrially relevant enzymes and is particularly known for the production of cellulases. Despite the availability of advanced gene editing techniques for T. reesei, the cultivation and characterization of resulting strain libraries remain challenging, necessitating well-defined and controlled conditions with higher throughput. Small-scale cultivation devices are popular for screening bacterial strain libraries. However, their current use for filamentous fungi is limited due to their complex morphology.ResultsThis study addresses this research gap through the development of a batch cultivation protocol using a microbioreactor for cellulase-producing T. reesei strains (wild type, RutC30 and RutC30 TR3158) with offline cellulase activity analysis. Additionally, the feasibility of a microscale fed-batch cultivation workflow is explored, crucial for mimicking industrial cellulase production conditions. A batch cultivation protocol was developed and validated using the BioLector microbioreactor, a Round Well Plate, adapted medium and a shaking frequency of 1000 rpm. A strong correlation between scattered light intensity and cell dry weight underscores the reliability of this method in reflecting fungal biomass formation, even in the context of complex fungal morphology. Building on the batch results, a fed-batch strategy was established for T. reesei RutC30. Starting with a glucose concentration of 2.5 g l-1\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$^{-1}$$\\end{document} in the batch phase, we introduced a dual-purpose lactose feed to induce cellulase production and prevent carbon catabolite repression. Investigating lactose feeding rates from 0.3 to 0.75 g (l h)-1\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$^{-1}$$\\end{document}, the lowest rate of 0.3 g (l h)-1\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$^{-1}$$\\end{document} revealed a threefold increase in cellobiohydrolase and a fivefold increase in β\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\upbeta$$\\end{document}-glucosidase activity compared to batch processes using the same type and amount of carbon sources.ConclusionWe successfully established a robust microbioreactor batch cultivation protocol for T. reesei wild type, RutC30 and RutC30 TR3158, overcoming challenges associated with complex fungal morphologies. The study highlights the effectiveness of microbioreactor workflows in optimizing cellulase production with T. reesei, providing a valuable tool for simultaneous assessment of critical bioprocess parameters and facilitating efficient strain screening. The findings underscore the potential of microscale fed-batch strategies for enhancing enzyme production capabilities, revealing insights for future industrial applications in biotechnology.

Performance evaluation of Trichoderma reseei in tolerance and biodegradation of diuron herbicide in agar plate, liquid culture and solid-state fermentation.

The present study evaluated the performance of the fungus Trichoderma reesei to tolerate and biodegrade the herbicide diuron in its agrochemical presentation in agar plates, liquid culture, and solid-state fermentation. The tolerance of T. reesei to diuron was characterized through a non-competitive inhibition model of the fungal radial growth on the PDA agar plate and growth in liquid culture with glucose and ammonium nitrate, showing a higher tolerance to diuron on the PDA agar plate (inhibition constant 98.63 mg L-1) than in liquid culture (inhibition constant 39.4 mg L-1). Diuron biodegradation by T. reesei was characterized through model inhibition by the substrate on agar plate and liquid culture. In liquid culture, the fungus biotransformed diuron into 3,4-dichloroaniline using the amide group from the diuron structure as a carbon and nitrogen source, yielding 0.154mg of biomass per mg of diuron. A mixture of barley straw and agrolite was used as the support and substrate for solid-state fermentation. The diuron removal percentage in solid-state fermentation was fitted by non-multiple linear regression to a parabolic surface response model and reached the higher removal (97.26%) with a specific aeration rate of 1.0 vkgm and inoculum of 2.6 × 108 spores g-1. The diuron removal in solid-state fermentation by sorption on barley straw and agrolite was discarded compared to the removal magnitude of the biosorption and biodegradation mechanisms of Trichoderma reesei. The findings in this work about the tolerance and capability of Trichoderma reesei to remove diuron in liquid and solid culture media demonstrate the potential of the fungus to be implemented in bioremediation technologies of herbicide-polluted sites.

Specific activity and utility of recombinant cellobiohydrolase II (Cel6A) produced in maize endosperm.

Cellobiohydrolase II (CBH II) is an exo-glucanase that is part of a fungal mixture of enzymes from a wood-rot fungus, Trichoderma reesei. It is therefore difficult to purify and to establish a specific activity assay. The gene for this enzyme, driven by the rice Os glutelin promoter, was transformed into High II tissue culture competent corn, and the enzyme accumulated in the endosperm of the seed. The transgenic line recovered from tissue culture was bred into male and female elite Stine inbred corn lines, stiff stalk 16083-025 (female) and Lancaster MSO411 (male), for future production in their hybrid. The enzyme increases its accumulation throughout its 6 generations of back crosses, 27-266-fold between T1 and T2, and 2-10-fold between T2 and T3 generations with lesser increases in T4-T6. The germplasm of the inbred lines replaces the tissue culture corn variety germplasm with each generation, with the ultimate goal of producing a high-yielding hybrid with the transgene. The CBH II enzyme was purified from T5 inbred male grain 10-fold to homogeneity with 47.5% recovery. The specific activity was determined to be 1.544 units per µg protein. The corn-derived CBH II works in biopolishing of cotton by removing surface fibers to improve dyeability and increasing glucose from corn flour for increasing ethanol yield from starch-based first-generation processes.

Variation of taste and odor compounds in tea beverage after microbial fermentation by HPLC–MS, GC×GC–O–MS, GC–MS, and sensory evaluation

Tea beverages are widely welcomed by consumers, and their flavor quality is particularly important. In this study, different tea extracts were fermented by Trichoderma reesei (TR), Aspergillus niger (AN), and lactic acid bacteria (LAB) to obtain the tea beverages, which were characterized by sensory evaluation as well as instrumental analysis. Nonvolatile compounds in tea beverages were determined by high-performance liquid chromatography–mass spectrometry (HPLC–MS), and volatile compounds were determined by solid-phase microextraction (SPME) combined with comprehensive two-dimensional gas chromatography–olfactory–mass spectrometry (GC×GC–O–MS) and gas chromatography–mass spectrometry (GC–MS). The data were analyzed by both principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA). The bitter and astringent galloylated catechins in tea beverages were hydrolyzed after TR and AN fermentation. Then after fermentation with LAB, some organic acid and phenolic compounds were decarboxylated, and some saccharides were also metabolized. Meanwhile, the increased content of alcohols and ketones in tea beverages affected the overall aroma. These results demonstrate the positive effects of microbial fermentation on the processing of tea beverages and provide technique support for comprehensive flavor quality control.