Termite Fungus Comb Polysaccharides Alleviate Hyperglycemia and Hyperlipidemia in Type 2 Diabetic Mice by Regulating Hepatic Glucose/Lipid Metabolism and the Gut Microbiota.

Type 2 diabetes (T2D) is a chronic metabolic disorder characterized by hyperglycemia and dyslipidemia. The termite fungus comb is an integral component of nests of termites, which are a global pest. Termite fungus comb polysaccharides (TFCPs) have been identified to possess antioxidant, anti-aging, and immune-enhancing properties. However, their physicochemical characteristics and their role in fighting diabetes have not been previously reported. In the current study, TFCPs were isolated and structurally characterized. The yield of TFCPs was determined to be 2.76%, and it was found to be composed of a diverse array of polysaccharides with varying molecular weights. The hypoglycemic and hypolipidemic effects of TFCPs, as well as their potential mechanisms of action, were investigated in a T2D mouse model. The results demonstrated that oral administration of TFCPs could alleviate fasting blood glucose levels, insulin resistance, hyperlipidemia, and the dysfunction of pancreatic islets in T2D mice. In terms of mechanisms, the TFCPs enhanced hepatic glycogenesis and glycolysis while inhibiting gluconeogenesis. Additionally, the TFCPs suppressed hepatic de novo lipogenesis and promoted fatty acid oxidation. Furthermore, the TFCPs altered the composition of the gut microbiota in the T2D mice, increasing the abundance of beneficial bacteria such as Allobaculum and Faecalibaculum, while reducing the levels of pathogens like Mailhella and Acetatifactor. Overall, these findings suggest that TFCPs may exert anti-diabetic effects by regulating hepatic glucose and lipid metabolism and the composition of the gut microbiota. These findings suggest that TFCPs can be used as a promising functional ingredient for the prevention and treatment of T2D.

γ-Glutamyl-ß-phenylethylamine, a novel α-glucosidase and α-amylase inhibitory compound from Termitomyces robustus, an edible Nigerian mushroom.

Termitomyces species are known edible mushrooms in Nigeria, believed to have exceptional culinary and nutraceutical properties. Methanol extract from fruiting bodies of Termitomyces robustus was evaluated for antidiabetic activity using in vitro α-amylase and α-glucosidase assays. The isolation and structural elucidation of metabolites from the T. robustus extract afforded five compounds including a new natural product γ-glutamyl-ß-phenylethylamine 3 and four known phenyl derivatives: tryptophan 1, 4-hydroxyphenylacetic acid 2, 4-hydroxyphenylpropionic acid 4, and phenyllactic acid 5. Structures were elucidated from analyses of spectroscopic data (1 D and 2 D NMR, HRESIMS) and all isolated compounds were tested for α-amylase and α-glycosidase inhibitory activity. The in vitro assay established crude extract to possess α- amylase and α-glucosidase inhibition with IC50 of 78.05 µg/mL and 86.10 µg/mL, respectively. The isolated compounds compared favourably with the standard drug, acarbose with IC50 ranging from 6.18-15.08 µg/mL and 18.28-44.63 µg/mL for α-amylase and glucosidase, respectively.

The Termite Fungal Cultivar Termitomyces Combines Diverse Enzymes and Oxidative Reactions for Plant Biomass Conversion.

Macrotermitine termites have domesticated fungi in the genus Termitomyces as their primary food source using predigested plant biomass. To access the full nutritional value of lignin-enriched plant biomass, the termite-fungus symbiosis requires the depolymerization of this complex phenolic polymer. While most previous work suggests that lignocellulose degradation is accomplished predominantly by the fungal cultivar, our current understanding of the underlying biomolecular mechanisms remains rudimentary. Here, we provide conclusive omics and activity-based evidence that Termitomyces employs not only a broad array of carbohydrate-active enzymes (CAZymes) but also a restricted set of oxidizing enzymes (manganese peroxidase, dye decolorization peroxidase, an unspecific peroxygenase, laccases, and aryl-alcohol oxidases) and Fenton chemistry for biomass degradation. We propose for the first time that Termitomyces induces hydroquinone-mediated Fenton chemistry (Fe2+ + H2O2 + H+ → Fe3+ + •OH + H2O) using a herein newly described 2-methoxy-1,4-dihydroxybenzene (2-MH2Q, compound 19)-based electron shuttle system to complement the enzymatic degradation pathways. This study provides a comprehensive depiction of how efficient biomass degradation by means of this ancient insect's agricultural symbiosis is accomplished. IMPORTANCE Fungus-growing termites have optimized the decomposition of recalcitrant plant biomass to access valuable nutrients by engaging in a tripartite symbiosis with complementary contributions from a fungal mutualist and a codiversified gut microbiome. This complex symbiotic interplay makes them one of the most successful and important decomposers for carbon cycling in Old World ecosystems. To date, most research has focused on the enzymatic contributions of microbial partners to carbohydrate decomposition. Here, we provide genomic, transcriptomic, and enzymatic evidence that Termitomyces also employs redox mechanisms, including diverse ligninolytic enzymes and a Fenton chemistry-based hydroquinone-catalyzed lignin degradation mechanism, to break down lignin-rich plant material. Insights into these efficient decomposition mechanisms reveal new sources of efficient ligninolytic agents applicable for energy generation from renewable sources.

Termitomyces heimii Associated with Fungus-Growing Termite Produces Volatile Organic Compounds (VOCs) and Lignocellulose-Degrading Enzymes.

Termitomyces fungi associated with fungus-growing termites are the edible mushrooms and can produce useful chemicals, enzymes, and volatile organic compounds (VOCs) that have both fuel and biological potentials. To this purpose, we examined the Termitomyces mycelial growth performance on various substrates, clarified lignocellulose-degrading enzyme activity, and also identified the VOCs produced by Termitomyces. Our results indicated that the optimal nutrition and condition requirements for mycelial growth are D-sorbitol, D-(+)-glucose, and D-(-)-fructose as carbon sources; peptone as well as yeast extract and ammonium tartrate as nitrogen sources; and Mn2+, Na+, and Mg2+ as metal ions with pH range from 7.0 to 8.0. Besides, the orthogonal matrix method results revealed that the ideal composition for mycelial growth is 20 g/L D-(-)-fructose, 5 g/L yeast extract, 0.5 g/L Mg2+, and pH = 7. We also screened various substrates composition for the activity of lignocellulose-degrading enzymes, i.e., lignin peroxidase, manganese peroxidase, ß-glucosidase, a-L-arabinofuranosidase, and laccase. Furthermore, we identified 37 VOCs using GC-MS, and the most striking aspect was the presence of a big series of alcohols and acids, collectively constituted about 49% of the total VOCs. Ergosta-5, 8, 22-trien-3-ol, (3.beta.,22E) was the most plenteous compound constituted 30.369%. This study hopes to establish a better understanding for researchers regarding Termitomyces heimii cultivation on a large scale for the production of lignocellulosic enzymes and some fungal medicine.

Anti-Diabetic Nephropathy Activities of Polysaccharides Obtained from Termitornyces albuminosus via Regulation of NF-κB Signaling in db/db Mice.

Termitornyces albuminosus is a kind of traditional Chinese edible fungus rich in nutrients and medicinal ingredients, and it has anti-oxidative, analgesic and anti-inflammatory effects. However, the hypoglycemic and nephroprotective effects of polysaccharides separated from T. albuminosus (PTA) have not been reported. The properties of PTA were analyzed in a BKS.Cg-Dock7m +/+ Leprdb/JNju (db/db) mouse model of diabetes. After the administration of PTA for eight weeks, the hypoglycemic and hypolipidemic activities of PTA in the db/db mice were assessed. The results of a cytokine array combined with an enzyme-linked immunosorbent assay confirmed the anti-oxidative and anti-inflammatory activities of PTA. An eight-week administration of PTA caused hypoglycemic and hypolipidemic functioning, as indicated by suppressed plasma glucose levels, as well as the modulation of several cytokines related to glycometabolism, in the sera and kidneys of the mice. PTA treatment also had a protective effect on renal function, restoring renal structures and regulating potential indicators of nephropathy. In the kidneys of the db/db mice, PTA treatment reduced the activation of protein kinase B, the inhibitor of κB kinase alpha and beta, and the inhibitor of κB alpha and nuclear factor-κB (NF-κB). We establish the hypoglycemic, hypolipidemic, and anti-diabetic nephropathy effects of PTA, and we find that the renal protection effects of PTA may be related to anti-inflammatory activity via the regulation of NF-κB signaling.

New Sesquiterpenoids from the Fermented Broth of Termitomyces albuminosus and their Anti-Acetylcholinesterase Activity.

Termitomyces albuminosus is the symbiotic edible mushroom of termites and cannot be artificially cultivated at present. In the project of exploring its pharmaceutical metabolites by microbial fermentation, four new selinane type sesquiterpenoids-teucdiol C (1), D (2), E (3), and F (4), together with two known sesquiterpenoids teucdiol B (5) and epi-guaidiol A (6)-were obtained from its fermented broth of T. albuminosus. Their structures were elucidated by the analysis of NMR data, HR Q-TOF MS spectral data, CD, IR, UV, and single crystal X-ray diffraction. Epi-guaidiol A showed obvious anti-acetylcholinesterase activity in a dose-dependent manner. The experimental results displayed that T. albuminosus possess the pharmaceutical potential for Alzheimer's disease, and it was an effective way to dig new pharmaceutical agent of T. albuminosus with the microbial fermentation technique.

Enzyme Activities at Different Stages of Plant Biomass Decomposition in Three Species of Fungus-Growing Termites.

Fungus-growing termites rely on mutualistic fungi of the genus Termitomyces and gut microbes for plant biomass degradation. Due to a certain degree of symbiont complementarity, this tripartite symbiosis has evolved as a complex bioreactor, enabling decomposition of nearly any plant polymer, likely contributing to the success of the termites as one of the main plant decomposers in the Old World. In this study, we evaluated which plant polymers are decomposed and which enzymes are active during the decomposition process in two major genera of fungus-growing termites. We found a diversity of active enzymes at different stages of decomposition and a consistent decrease in plant components during the decomposition process. Furthermore, our findings are consistent with the hypothesis that termites transport enzymes from the older mature parts of the fungus comb through young worker guts to freshly inoculated plant substrate. However, preliminary fungal RNA sequencing (RNA-seq) analyses suggest that this likely transport is supplemented with enzymes produced in situ Our findings support that the maintenance of an external fungus comb, inoculated with an optimal mixture of plant material, fungal spores, and enzymes, is likely the key to the extraordinarily efficient plant decomposition in fungus-growing termites.IMPORTANCE Fungus-growing termites have a substantial ecological footprint in the Old World (sub)tropics due to their ability to decompose dead plant material. Through the establishment of an elaborate plant biomass inoculation strategy and through fungal and bacterial enzyme contributions, this farming symbiosis has become an efficient and versatile aerobic bioreactor for plant substrate conversion. Since little is known about what enzymes are expressed and where they are active at different stages of the decomposition process, we used enzyme assays, transcriptomics, and plant content measurements to shed light on how this decomposition of plant substrate is so effectively accomplished.

Complementary symbiont contributions to plant decomposition in a fungus-farming termite.

Termites normally rely on gut symbionts to decompose organic matter but the Macrotermitinae domesticated Termitomyces fungi to produce their own food. This transition was accompanied by a shift in the composition of the gut microbiota, but the complementary roles of these bacteria in the symbiosis have remained enigmatic. We obtained high-quality annotated draft genomes of the termite Macrotermes natalensis, its Termitomyces symbiont, and gut metagenomes from workers, soldiers, and a queen. We show that members from 111 of the 128 known glycoside hydrolase families are represented in the symbiosis, that Termitomyces has the genomic capacity to handle complex carbohydrates, and that worker gut microbes primarily contribute enzymes for final digestion of oligosaccharides. This apparent division of labor is consistent with the Macrotermes gut microbes being most important during the second passage of comb material through the termite gut, after a first gut passage where the crude plant substrate is inoculated with Termitomyces asexual spores so that initial fungal growth and polysaccharide decomposition can proceed with high efficiency. Complex conversion of biomass in termite mounds thus appears to be mainly accomplished by complementary cooperation between a domesticated fungal monoculture and a specialized bacterial community. In sharp contrast, the gut microbiota of the queen had highly reduced plant decomposition potential, suggesting that mature reproductives digest fungal material provided by workers rather than plant substrate.

Tamarind kernel powder: a novel agro-residue for the production of cellobiose dehydrogenase under submerged fermentation by Termitomyces clypeatus.

The study investigates the potential of substitution of the conventional carbohydrate nutrient (cellulose) in media with cheap agro-residues for cellobiose dehydrogenase production by Termitomyces clypeatus (CDHtc) under submerged conditions. Different agro-residues tested for enzyme production were characterized using FTIR and XRD analysis. As CDHtc production was highest with tamarind kernel powder (TKP), it was selected for process optimizations through shake-flask fermentations. The optimized parameters were then applied to batch cultures in a 5 L bioreactor that gave enzyme yield (57.4 U mL⁻¹) similar to that obtained under shake-flask fermentations (57.05 U mL⁻¹). The study also made an attempt to predict CDHtc production with respect to time of fermentation and mycelial growth. The specific growth rate and carrying capacity of the mycelia were also determined, and the values lie in the ranges of 0.024-0.027 h⁻¹ and 7.2-7.1 mg mL⁻¹, respectively.

Synergistic collaboration of gut symbionts in Odontotermes formosanus for lignocellulosic degradation and bio-hydrogen production.

In this work, gut microbes from the macrotermitine termite Odontotermes formosanus the cellulolytic Bacillus and fermentative Clostridium were studied in batch experiments using different carbon substrates to bio-mimic the termite gut for hydrogen production. Their fungus comb aging and the in vitro lignocellulosic degradation of the mango tree substrates by the synergistic interaction of Bacillus, Clostridium and Termitomyces were detected by Solid-state NMR. From the results, Bacillus species acted as a mutualist, by initiating an anaerobic environment for the growth of Clostridium, for bio-hydrogen production and the presence of Termitomyces enhanced the lignocellulosic degradation of substrates in vitro and in vivo. Thus, the synergistic collaboration of these three microbes can be used for termite-derived bio-fuel processing technology.

Transcriptome analysis of Termitomyces albuminosus reveals the biodegradation of lignocellulose.

OBJECTIVE: To study whether Termitomyces albuminosus can degrade lignocelluloses and to understand the symbiotic relationship between termite mushroom and fungus-growing termite. METHODS: cDNA library of T. albuminosus was sequenced by the Roche 454 GS FLX Titanium platform, and the diverse enzymes relevant to degradation of cellulose and lignin of symbiotic fungus T. albuminosus were analyzed. RESULTS: Eighth sequencing run resulted in a total of 82386 reads (express sequence tags, EST). After removing the vector and primer sequences, the remained 54410 reads were assembled into 3301 contigs and 3193 singletons. Comparing sequence similarity with known proteins, these sequences, representing approximately 2681 unique genes, were successfully annotated using BLAST searches (E-value < or = 1e(-10)) against the Nr, SwissProt and CDD databases. The T. albuminosus transcripts included 33 enzymes putatively involved in cellulose and hemicelluloses biodegradation. 5 enzymes could hydrolyze cellulose and others had catalytic activities for degradation of hemicelluloses, starch and glycogen and chitin. Moreover, four genes encoding laccases and a single aryl-alcohol oxidase which could degrade lignin were also identified. These results revealed symbiosis fungus T. albuminosus had many laccases and possibly decomposed phenolic compounds from plant litter. CONCLUSIONS: Data presented in this study indicated that T. albuminosus had the ability to degrade lignin, which made cellulose more easily degraded by the cellulase produced by fungus-growing termite.

De novo sequencing and analysis of the termite mushroom (Termitomyces albuminosus) transcriptome to discover putative genes involved in bioactive component biosynthesis.

A one-eighth 454 sequencing run produced 82,386 high-quality reads. De novo assembly generated 6494 unique sequences. Based on the bioinformatic analysis, we found many the known enzymes involved in the biosynthesis of triterpene saponin in Termitomyces albuminosus, including 6 cytochrome P450 and 22 glycosyltransferase unique genes.

Increased enzyme secretion by 2-deoxy-D-glucose in presence of succinate by suppression of metabolic enzymes in Termitomyces clypeatus.

Regulatory mode of secretion of proteins was detected for the industrial glycosidase, cellobiase, under secreting conditions (in presence of TCA cycle intermediates like succinate etc.) in the filamentous fungus Termitomyces clypeatus. The titers of key metabolic enzymes were investigated under secreting and non-secreting conditions of growth and compared to the corresponding production of intra and extracellular levels of cellobiase. Results were compared in presence of 2-deoxy-D-glucose, a potent glycosylation inhibitor in the secreting media. Inclusion of 2-deoxy-D-glucose in presence of succinate caused about 10 to 100 times decrease in titers of the metabolic enzymes hexokinase, fructose-1,6-bisphosphatase, isocitrate lyase and malate dehydrogenase leading to increased secretion of cellobiase by more than 100 times. The intracellular concentration of cAMP (86-fold decrease in presence of 2-deoxy-D-glucose under secreting conditions) and turnover rate of proteins also dropped significantly. In this suppressed metabolic state, a 10-fold increase in the titer of the secreted cellobiase was noticed. The results indicated elucidation of carbon catabolite repression like phenomenon in the fungus under secreting conditions which was more pronounced by 2-deoxy-D-glucose. The interdependence between secretion and regulation of metabolic enzymes will help in better understanding of the physiology of these highly adapted organisms for increasing their secretion potential of glycosidases like cellobiase with high industrial value.