Overexpression of cat2 restores antioxidant properties and production traits in degenerated strains of Volvariella volvacea.

Strain degeneration is an important factor hindering the development of the edible fungus industry. Strain degeneration is associated with the excessive accumulation of reactive oxygen species (ROS) in vivo. Catalase (CAT), an important antioxidant enzyme, can promote the clearance of ROS. In this study, the cat2 gene of Volvariella volvacea was first cloned into an overexpression plasmid via homologous recombination. Finally, through Agrobacterium-mediated transformation, this plasmid was inserted into degenerated strains of V. volvacea T19. The physiological properties, antioxidant properties, ROS content, matrix degradation activity, and cultivation properties of the transformants were tested. The results showed that the cloned cat2 gene was 99.94% similar to the reference sequence. Screening revealed that six positive transformants were successfully obtained. After the overexpression of cat2, the growth rate and biomass of the mycelium increased significantly in the transformant strains (versus the V. volvacea T19 degenerated strains). Moreover, the accumulation of superoxide radical (O2•-) and hydrogen peroxide (H2O2) was significantly reduced, and the activity of the enzymes CAT, superoxide dismutase (SOD), glutathione reductase (GR), and glutathione peroxidase (GPX) was significantly increased. Meanwhile, the expression of cat2, Mnsod1, Mnsod2, gpx, and gr was significantly upregulated, and the activity of eight matrix degradation-related enzymes was increased to varying degrees. More importantly, the overexpression of the cat2 gene promoted the regrowth of fruiting bodies in degenerated strains of V. volvacea T19. This study provides a new biotechnological strategy to control the degeneration of V. volvacea and other edible fungi.

The combination of high oxygen and nanocomposite packaging alleviated quality deterioration by promoting antioxidant capacity and phenylpropane metabolism in Volvariella volvacea.

Volvariella volvacea is a highly perishable mushroom that severely affects its postharvest commercial value. This study aimed to investigate the impact of high oxygen (O2) levels combined with nanocomposite packaging on the shelf-life quality of V. volvacea. Results showed that treatment with high concentrations of O2 (80% and 100% O2) and nanocomposite packaging effectively delayed the quality deterioration of V. volvacea, resulting in better postharvest appearance, higher firmness, lower weight loss, malondialdehyde (MDA) content, and leakage of membrane electrolytes. Further analysis revealed the combination treatments ameliorated oxidative stress by inducing antioxidant enzymes and the glutathione-ascorbate (GSH-AsA) cycle at both enzymatic and transcriptional levels, thereby activating the antioxidant system. Additionally, the treatments enhanced activities of key enzymes in phenylpropane metabolism, leading to a reduction in the decrease of total phenolics and flavonoids. This work provides new insights into the development of postharvest technologies to prolong the storage life of V. volvacea.

Volvariella volvacea brain Abscess in an immunocompromised host-An emerging fungal pathogen in Asia.

Volvariella volvacea is a fungus found in tropical regions, commonly associated with straw mushrooms. This is a 50-year-old Singaporean female post living donor renal transplant who presented with fever, cough and headache. She was diagnosed to have Volvariella volvacea brain abscess. She was treated with combination anti-fungal therapy without surgical debridement and remains stable. The pathogenicity of this rare fungus in immunocompromised hosts is demonstrated here and is of significance particularly in Asia where ingestion of straw mushrooms may be a risk factor for invasive fungal disease.

Bioconversion of bamboo shoot shells through the cultivation of the edible mushrooms Volvariella volvacea.

Bamboo shoot shell (BSS), as agricultural waste, is mostly burned or discarded, causing serious environment pollution. In this study, the degradation and utilization of BSS by the edible fungus Volvariella Volvacea was investigated. The composition of V. volvacea fruit body was determined by HPLC-MS, GC-MS and ICP-OES. The activities of CMCase and xylanase were monitored by DNS (3,5-dinitrosalicylic acid) method. Laccase activity was assayed by the oxidation reaction of ABTS [2,2'-azinobis-(3-ethylbenzthiazoline-6-sulphonate)]. The degraded bamboo shoot shell powder was characterized by FTIR and SEM. The results showed that the mycelium of V. volvacea could degrade and utilize BSS for growth. The activities of carboxymethyl cellulase and laccase were increased during the cultivation. At the same time, the physical structure of the shell fiber becames porous and rough. Most of the products of decayed fibers contain alkanes, ethyl or methyl groups. Moreover, the biological efficiency (fruiting body yield) of V. volvacea cultivated on BSS was 1.52-fold higher than that of straw cultivation. The contents of total lipid, elaidic acid (C18:1n-9), total essential amino acids, total amino acids and iron in V. volvacea fruit bodies grown on BSS were 1.11, 1.66, 1.52, 1.60 and 1.30-fold higher than those of straw treatment, respectively. This study provides an effective method to solve the environmental pollution caused by BSS, and provides a new way for the potential utilization of BSS in edible fungi cultivation.

Volvariella turcica, a new species from Turkey, and a multigene phylogeny of Volvariella.

A new species of Volvariella, collected from Aydin Province on the coast of the Aegean Sea in southwestern Turkey, is described as Volvariella turcica, sp. nov., based on morphology and multigene molecular analysis of three nuc rDNA gene regions: internal transcribed spacer ITS1-5.8S-ITS2 (ITS), 28S, and 18S. The new species was found in forests dominated by Pinus brutia and Quercus coccifera and mainly characterized by small basidiomata with a white pileus covered with pale ochre center and an ochre-discoloring volva, small basidiospores, lageniform pleurocystidia, balloon-shaped to clavate cheilocystidia, and stipitipellis hairs that are cylindrical or cylindrical-tortuous with subcapitate or lobe-like projections. A comprehensive description, illustrations, and line drawings are provided, and comparison with morphologically similar and phylogenetically related species is discussed.

A Comparative Transcriptome Analysis of Volvariella volvacea Identified the Candidate Genes Involved in Fast Growth at the Mycelial Growth Stage.

The edible straw mushroom, Volvariella volvacea, is one of the most important cultivated mushrooms in tropical and sub-tropical regions. Strain improvement for V. volvacea is difficult because of the unknown mechanisms involved in its growth regulation and substrate utilization. A comparative physiological and transcriptomic study was conducted between two commercially available straw mushroom strains (v9 and v26) to explore their fast-growth regulation mechanism(s). The physiological study showed that V. volvacea v9 had a shorter growth cycle and higher biological efficiency (4% higher) than that in v26. At least 14,556 unigenes were obtained from the four cDNA libraries (two replicates per strain). Among them, the expression of 1597 unigenes was up-regulated while 1352 were down-regulated. Four heat-shock proteins were highly expressed in v9, showing that v9 has the better ability to handle stresses and/or environmental changes. Moreover, up to 14 putative transporter genes were expressed at a higher level in v9 than those in v26, implying that v9 has a better ability to transport nutrients or export xenobiotics efficiently. Our report allows to identify the candidate genes involved in the fast growth requirement of V. volvacea, which represents a valuable resource for strain improvement in this commercially important edible mushroom.

A macromolecular α-glucan from fruiting bodies of Volvariella volvacea activating RAW264. 7 macrophages through MAPKs pathway.

A novel macromolecular polysaccharide VGPⅠ-a was purified from Volvariella volvacea fruiting bodies with ultrasound-assisted extraction, ion exchange and gel chromatography. VGPⅠ-a was proved as a α- glucan with Mw of 1435.6 kDa and contained a 1,4-linked d-Glcp backbone with the substitution at C-6 with 1-linked d-Glcp residue. Congo-red test, AFM and SEM analysis showed VGPⅠ-a had a triple-helical conformation and the interacted chains to form a small screw-rod and dispersed appearance. VGPⅠ-a had no cytotoxic effect on macrophage RAW264.7 cells in vitro and significantly enhanced the production and mRNA expression of NO, TNF-α, IL-6 and IL-1ß in a dose-dependent manner. Further analyses demonstrated that VGPⅠ-a activated the MAPK signaling pathway by improving the phosphorylated levels of p38, JNK and ERK in RAW264.7 cells to promote the expression and secretion of above cytokines. These findings would provide a better understanding of V. volvacea glucan and its potential immunomodulating mechanisms.

Chilling stress reduced protein translation by the ubiquitination of ribosomal proteins in Volvariella volvacea.

In Volvariella volvacea, an important edible mushroom species, cryogenic autolysis is a typical part of abnormal metabolism; however, the underlying mechanisms remain unclear. Ubiquitylome analysis revealed that chilling stress (CS) affected protein translation and degradation by ubiquitination. Comparative proteomics analysis showed that CS downregulated protein expression in V. volvacea V23 instead of VH3 (improved chilling stress resistance strain). The integrative ubiquitylome, proteomics, and transcriptome analyses indicated that CS reduced protein translation by the ubiquitination of ribosomal proteins. An activity assay of the 20S proteasome showed that CS decreased the degradation efficiency of the ubiquitin-proteasome system. UBEV2, one type of ubiquitin-conjugating enzyme E2 (UBE2) in V. volvacea, was upregulated after cold stress treatment using western blot analysis. GST pull-down experiments of UBEV2 provided evidence that CS affected protein translation by the ubiquitination of ribosomal proteins. Co-IP experiments confirmed that UBEV2 bound to the ubiquitinated SSB2, a ribosome-associated molecular chaperone. An anti-freezing experiment demonstrated that the UBE2 inhibitor could improve the cold stress resistance of V. volvacea. Our observations revealed that CS triggered ubiquitination-mediated autolysis associated with a decrease in protein translation and highlighted the mechanistic role of UBEV2 in facilitating cryogenic autolysis in V. volvacea. SIGNIFICANCE: Volvariella volvacea, the edible straw mushroom, is a highly nutritious food source widely cultivated on a commercial scale in tropical and subtropical regions. The challenges associated with the cryogenic autolysis preservation of V. volvacea have limited its marketability. This issue of cryogenic autolysis is both an interesting scientific problem to solve and a practical economic matter. Integrative ubiquitylome, proteomics, and transcriptome analyses, together with GST pulldown and Co-IP experiments, indicated that chilling stress reduced protein translation by the ubiquitination of ribosomal proteins in V. volvacea. This study significantly contributes to our understanding of ubiquitination-mediated autolysis associated with a decrease in protein translation in V. volvacea. Our data highlight the mechanistic role of UBEV2 in facilitating the cryogenic autolysis of V. volvacea. We provided a new idea for the preservation of V. volvacea by inhibiting UBEV2 to increase its marketability.

Study on chemical constituents of an edible mushroom Volvariella volvacea and their antitumor activity in vitro.

Phytochemical investigation of the fruiting body of Volvariella volvacea led to the isolation of a new furanone, 2(5H)-furanone-4-propionic acid named volvafuranone A (1), together with twelve known compounds (2-13). Compounds 2-7, 9-11 were isolated from this mushroom for the first time. The isolated compounds were assessed for their cytotoxicity against four human tumour lines (SGC-7901, PC-3M, MCF-7, HepG-2), and the results showed that compound 2, 3, 12, 13 have significant cytotoxicity with IC50 values of 5.90 µM (HepG-2), 20.72 µM (HepG-2), 27.98 µM (PC-3M) and 23.15 µM (PC-3M), respectively.

The NADPH oxidase in Volvariella volvacea and its differential expression in response to mycelial ageing and mechanical injury.

NADPH oxidases are enzymes that have been reported to generate reactive oxygen species (ROS) in animals, plants and many multicellular fungi in response to environmental stresses. Six genes of the NADPH oxidase complex components, including vvnoxa, vvnoxb, vvnoxr, vvbema, vvrac1 and vvcdc24, were identified based on the complete genomic sequence of the edible fungus Volvariella volvacea. The number of vvnoxa, vvrac1, vvbema and vvcdc24 transcripts fluctuated with ageing, and the gene expression patterns of vvnoxa, vvrac1 and vvbema were significantly positively correlated. However, the expression of vvnoxb and vvnoxr showed no significant difference during ageing. In hyphae subjected to mechanical injury stress, both O2- and H2O2 concentrations were increased. The expression of vvnoxa, vvrac1, vvbema and vvcdc24 was substantially upregulated, but vvnoxb and vvnoxr showed no response to mechanical injury stress at the transcriptional level. Additionally, the transcription of vvnoxa, vvrac1, vvbema and vvcdc24 could be repressed when the intracellular ROS were eliminated by diphenyleneiodonium (DPI) chloride and reduced glutathione (GSH) treatments. These results indicated a positive feedback loop involving NADPH oxidase and intracellular ROS, which might be the reason for the oxidative burst during injury stress.

Changes in Mannitol Content, Regulation of Genes Involved in Mannitol Metabolism, and the Protective Effect of Mannitol on Volvariella volvacea at Low Temperature.

The mechanism of autolysis of Volvariella volvacea (V. volvacea) at low temperature has not been fully explained. As mannitol is among the most important osmotic adjustment substances in fungal resistance, this study sampled mycelia of strains V23 and VH3 treated at 0°C for 0, 2, 4, 8, and 10 h to analyze changes in intracellular mannitol content by high-performance anion chromatography with pulsed amperometric detection (HAPEC-PAD). Reverse transcription quantitative PCR (RT-qPCR) analysis was applied to assess differences in the transcript levels of genes associated with mannitol metabolism under low-temperature stress. A mannitol solution was added to cultures of V. volvacea fruiting bodies, and effects on the hypothermic resistance of these organs were explored by evaluating variations in sensory properties during cryogenic storage after harvest. The results suggested that in the initial stage of low-temperature treatment, intracellular mannitol was largely catabolized as an energy storage material and the expression of genes encoding enzymes involved in synthetic reactions was inhibited. However, low-temperature resistance was induced with further treatment, with activation of mannitol synthesis and inhibition of degradation; the cells accumulated mannitol, leading to osmoregulation. No significant elongation of V. volvacea fruiting bodies during storage at 4°C was observed, and these organs tended to shrink and collapse. The sensory quality of mannitol-treated fruiting bodies was much better than that of control fruiting bodies. Application of a mannitol solution at the cultivation stage of V. volvacea somewhat improved the low-temperature resistance of the fruiting bodies, verifying the correlation between mannitol and resistance to this stress in V. volvacea. The results of this study lay a foundation for a deeper understanding of the autolysis mechanism of V. volvacea, providing technical support for increasing the cryopreservation time of this species and extending the postharvest shelf life of its fruiting bodies. In addition, the mechanism underlying the low-temperature tolerance of the VH3 strain should be further explained at the molecular level.

Identification of dihydro-ß-ionone as a key aroma compound in addition to C8 ketones and alcohols in Volvariella volvacea mushroom.

The volatile compounds of Volvariella volvacea mushroom were investigated by solvent assisted flavor evaporation (SAFE)/gas chromatography-mass spectrometry (GC-MS), gas chromatography-olfactometry (GC-O), odor activity value (OAV), combined with aroma reconstitution and omission. The results showed that a total of 63 compounds were detected after SAFE extraction. A total of 26 compounds were determined after GC-O and 17 compounds whose OAV greater than 1 were subjected to reconstitution and omission experiments. The results showed that dihydro-ß-ionone, 1-octen-3-one, 1-octen-3-ol, γ-undecalactone, 3-octanol, 2-octanone, hexanal, 2-methylbutanal, camphene, carvone, 2-nonanone, and phenylacetaldehyde have been successfully identified as the key aroma compounds. More significantly, dihydro-ß-ionone as a key aroma compound was first found in Volvariella volvacea mushroom.

Identification of umami-tasting peptides from Volvariella volvacea using ultra performance liquid chromatography quadrupole time-of-flight mass spectrometry and sensory-guided separation techniques.

Mushroom peptides produced by high-pressure cooking from Volvariella volvacea were found to retain and enhance the umami taste. Three tasty peptides were identified as Ala-Ser-Asn-Met-Ser-Asp-Leu (ASNMSDL), Tyr-Tyr-Gly-Ser-Asn-Ser-Ala (YYGSNSA) and Leu-Gln-Pro-Leu-Asn-Ala-His (LQPLNAH) by UPLC-Q-TOF-MS, after a series of separation methods of ultrafiltration, gel filtration chromatography and reverse phase high-performance liquid chromatography. Each fractionation step is based on the results of sensory evaluation. ASNMSDL and LQPLNAH have umami taste with threshold values of 10.19 and 12.63 mmol/L, respectively. In addition, these two peptides also showed umami-enhancing properties with a threshold estimated at 13.58 and 18.95 mmol/L, respectively. Furthermore, the peptides showed better sensory taste than mixtures of their constituent amino acids. After separation and purification, the content of taste peptides in Volvariella volvacea is about 1.4%.

Characterization and expression pattern of homeobox transcription factors in fruiting body development of straw mushroom Volvariella volvacea.

A number of homeobox transcription factors (TFs) play critical role in regulating developmental processes of fungi. However, studies on TFs in fruiting body development of mushroom forming species, Volvariella volvacea, are still at initial stage. Here, we report homeobox TFs in the whole-genomic sequence of V. volvacea and expression analysis of the homeobox TFs during a series of developmental stages. Homeobox TFs were identified using InterPro terms and Fungal Transcription Factor Database (FTFD) from the genome of V. volvacea and quantitative real-time PCR were used for gene expression analysis. Based on phylogenetic analysis, the homeobox TFs of V. volvacea were divided into two groups and showed close relationships with the TFs of other Basidiomycetes. Eight differentially expressed homeobox TFs were selected by digital gene expression analysis from 47 putative homeobox TFs, including five up-regulated genes in primordia and three down-regulated genes in fruiting elongation stage of V. volvacea. VvHox1, VvHox2, and VvHox3 might be participating in fruiting body elongation. It can be assumed that VvHox3 might be involved in volva development. Moreover, five TFs (VvHox4-VvHox8) might be contributing in primordia formation. Results indicated that differentially expressed homeobox TFs are significant candidates for fruiting body development study in V. volvacea.

Selection and Evaluation of Appropriate Reference Genes for RT-qPCR Normalization of Volvariella volvacea Gene Expression under Different Conditions.

Volvariella volvacea (V. volvacea), commonly referred to as Chinese (paddy straw) mushroom, is a basidiomycete with a protein-rich volva and pileus. Selecting appropriate reference genes is a crucial step in the normalization of quantitative real-time PCR data. Therefore, 12 candidate reference genes were selected from the V. volvacea transcriptome based on previous studies and then BestKeeper, geNorm, and NormFinder were used to identify reference genes stably expressed during different developmental stages and conditions. Of the 12 candidate reference genes, SPRY domain protein (SPRYp), alpha-tubulin (TUBα), cyclophilin (CYP), L-asparaginase (L-asp), and MSF1-domain-containing protein (MSF1) were the most stably expressed under different experimental conditions, while 18S ribosomal RNA (18S), 28S ribosomal RNA (28S), and beta-actin (ACTB) were the least stably expressed. This investigation not only revealed potential factors influencing the suitability of reference genes, but also identified optimal reference genes from a pool of candidate genes under a wide range of conditions.

Use of straw mushrooms (Volvariella volvacea) for the enhancement of physicochemical, nutritional and sensory profiles of Cantonese sausages.

The aim of this work was to evaluate the quality and sensory properties of Cantonese sausages incorporated with dried straw mushrooms. Five types of sausage with the addition of 0%, 1%, 2%, 3%, and 4% straw mushrooms were prepared, and their physicochemical traits and sensory acceptance were investigated. The results showed that incorporation of straw mushrooms improved the physical properties, significantly decreased peroxide value, increased the amount of amino acids and volatile compounds, and introduced new volatiles (aldehyde and ester compounds) to Cantonese sausages. Compared to sausages without addition of straw mushrooms, the amount of essential amino acids increased 8-fold, the P/S ratio of fatty acid increased to 0.46 similar to the recommended nutritional value, and the lipid peroxide value reduced 10-fold in Cantonese sausages with addition of 4% straw mushrooms. These results indicate that incorporation of straw mushrooms could be an efficient way to obtain nutritional and healthy Cantonese sausages.

Phenolics, tocopherols and fatty acid profiling of wild and commercial mushrooms from Pakistan.

Mushrooms can be used as nutraceutical or functional foods to maintain and promote good health. In the present study, wild Ganoderma lucidum and four commercial mushrooms, Pleurotus ostreatus, Volvariella volvacea, Hericium erinaceus and Lentinus edodes, collected from Pakistan were screened for phenolics, tocopherols and fatty acid contents. High performance liquid chromatography analysis of phenolic acids showed that chlorogenic acid, ferulic acid, gallic acid, p-Coumaric and caffeic acids were observed in selected mushrooms. H. erinaceus contained high amounts of chlorogenic acid (11.49±0.1 µ/g of dry weight) and ferulic acid (7.84±0.7 µg/g of dry weight). γ-tocopherol and lutein were present in all studied mushrooms. Lutein contents were higher in H. erinaceus (2.42±0.087 µg/g of DW) followed by V. volvacea> P. ostreatus> L. edodes. γ-tocopherol was observed in the range of 74.25±3.01 to 29.65±1.2 µg/g of dry weight. GC/MS analysis of fatty acids showed that linoleic acid (18’:2n6c), oleic acid (18’:1n9c), palmitic acid (C16:0), stearic acid (C18:0), linolenic acid (18’:3n3) and nonadecanoic acid (C19-0), were the main fatty acids found in selected mushrooms. The unsaturated fatty acids were predominated over saturated fatty acids. It is concluded that selected mushrooms are good sources of antioxidant compounds and unsaturated fatty acids.