Comparative genomics reveals ample evidence to Ganoderma sinense cultivars for molecular identification and new FIP exploration.

The first dikaryotic genome of Ganoderma cultivar Zizhi S2 (56.76 Mb, 16,681 genes) has been sequenced recently. 98.15% of complete BUSCOs were recovered in this genome assembly and high-confidence annotation rate improved to 91.41%. Collinearity analysis displayed the nuclear genome were 80.2% and 93.84% similar to reference genome of G. sinense at nucleotide and amino acid levels, which presented 8,521 core genes and 880 unique orthologous gene groups. Among that, at least six functional genes (tef1-α, ß-tubulin, rpb2, CaM, Mn-SOD and VeA) and a newly discovered fip gene were highly similar 99.27% ∼100% to those in reference genome. And the mt-LSU, mt-SSU and 13 PCGs in their mitogenome were also highly conserved with 99.27%-99.87% and 99.08%-100% identity, respectively. So that, this cultivar Zizhi S2 is confirmed conspecific with Ganoderma sinense (NCBI: txid1077348). The new fip gene (MN635280.1_336bp) existing a novel mutation which can be reflected on the phylogenetic tree and 3-dimensional model topology structure.

Insights into Ganoderma fungi meroterpenoids opening a new era of racemic natural products in mushrooms.

Ganoderma meroterpenoids (GMs) containing 688 structures to date were discovered to have multiple remarkable biological activities. 65.6% of meroterpenoids featuring stereogenic centers from Ganoderma species are racemates. Further, GMs from different Ganoderma species seem to have their own characteristics. In this review, a comprehensive summarization of GMs since 2000 is presented, including GM structures, structure corrections, biological activities, physicochemical properties, total synthesis, and proposed biosynthetic pathways. Additionally, we especially discuss the racemic nature, species-related structural distribution, and structure-activity relationship of GMs, which will provide a likely in-house database and shed light on future studies on GMs.

Total Synthesis of Nona-decasaccharide Motif from Ganoderma sinense Polysaccharide Enabled by Modular and One-Pot Stereoselective Glycosylation Strategy.

Polysaccharides from a medicinal fungus Ganoderma sinense represent important and adjunctive therapeutic agents for treating various diseases, including leucopenia and hematopoietic injury. However, the synthetic accessibility to long, branched, and complicated carbohydrates chains from Ganoderma sinense polysaccharides remains a challenging task in chemical synthesis. Here, we report the modular chemical synthesis of nona-decasaccharide motif from Ganoderma sinense polysaccharide GSPB70-S with diverse biological activities for the first time through one-pot stereoselective glycosylation strategy on the basis of glycosyl ortho-(1-phenyvinyl)benzoates, which not only sped up carbohydrates synthesis but also reduced chemical waste and avoided aglycones transfer issues inherent to one-pot glycosylation on the basis of thioglycosides. The synthetic route also highlights the following key steps: (1) preactivation-based one-pot glycosylation for highly stereoselective constructions of several 1,2-cis-glycosidic linkages, including three α-d-GlcN-(1 → 4) linkages and one α-d-Gal-(1 → 4) bond via the reagent N-methyl-N-phenylformamide modulation; (2) orthogonal one-pot assembly of 1,2-trans-glycosidic linkages in various linear and branched glycans fragments by strategic combinations of glycosyl N-phenyltrifluoroacetimidates, glycosyl ortho-alkynylbenzoates, and glycosyl ortho-(1-phenyvinyl)benzoates; and (3) the final [1 × 4 + 15] Yu glycosylation for efficient assembly of nona-decasaccharide target. Additionally, shorter sequences of 4-mer, 5-mer, and 6-mer are also prepared for structure-activity relationship biological studies. The present work shows that this one-pot stereoselective glycosylation strategy can offer a reliable and effective means to streamline chemical synthesis of long, branched, and complex carbohydrates with many 1,2-cis-glycosidic bonds.

1H NMR analysis of metabolites from leaf tissue of resistant and susceptible oil palm breeding materials against Ganoderma boninense.

INTRODUCTION: Breeding for oil palm resistance against basal stem rot caused by Ganoderma boninense is challenging and time-consuming. Advanced oil palm gene pools are very limited, hence it is assumed that parental palms have experienced genetic drift and lost their resistance genes against Ganoderma. High-throughput selection criteria should be developed. Metabolomic analysis using 1H nuclear magnetic resonance (NMR) spectroscopy is easy, and the resulting metabolite can be used as a diagnostic tool for detecting disease in various host-pathogen combinations. OBJECTIVES: The objective of this study was to identify metabolite variations in Dura (D) and Pisifera (P) parental palms with different resistance levels against Ganoderma and moderately resistant DxP using 1H NMR analysis. METHODS: Leaf tissues of seven different oil palm categories consisting of: resistant, moderate, and susceptible Dura (D); moderate and susceptible Pisifera (P); resistant Tenera/Pisifera (T/P) parental palms; and moderately resistant DxP variety progenies, were sampled and their metabolites were determined using NMR spectroscopy. RESULTS: Twenty-nine types of metabolites were identified, and most of the metabolites fall in the monosaccharides, amino acids, and fatty acids compound classes. The PCA, PLS-DA, and heatmap multivariate analysis indicated two identified groups of resistance based on their metabolites. The first group consisted of resistant T/P, moderate P, resistant D, and moderately resistant DxP. In contrast, the second group consisted of susceptible P, moderate D, and susceptible D. Glycerol and ascorbic acid were detected as biomarker candidates by OPLS-DA to differentiate moderately resistant DxP from susceptible D and P. The pathway analysis suggested that glycine, serine, and threonine metabolism and taurine and hypotaurine metabolism were involved in the oil palm defense mechanism against Ganoderma. CONCLUSION: A metabolomic study with 1H NMR was able to describe the metabolite composition that could differentiate the characteristics of oil palm resistance against basal stem rot (BSR) caused by G. boninense. These metabolites revealed in this study have enormous potential to become support tools for breeding new oil palm varieties with higher resistance against BSR.

Total Synthesis of Ganoderma Meroterpenoids Cochlearol B and Its Congeners Driven by Structural Similarity and Biological Homology.

Secondary metabolites that have the same biological origin must share some relationship in their biosynthesis. Exploring this relationship has always been a significant task for synthetic biologists. However, from the perspective of synthetic chemists, it is equally important to propose, prove, or refute potential biosynthetic pathways in order to elucidate and understand the biosynthesis of homologous secondary metabolites. In this study, driven by the high structural similarity between the homologous Ganoderma meroterpenoids cochlearol B and ganocin B, two chemically synthetic strategies were designed and investigated sequentially for the synthesis of cochlearol B from ganocin B. These strategies include intramolecular metal-catalyzed hydrogen atom transfer (MHAT) and intramolecular photochemical [2+2] cycloaddition. The aim was to reveal their potential biosynthetic conversion relationship using chemical synthesis methods. As a result, a highly efficient total synthesis of cochlearol B, cochlearol T, cochlearol F, as well as the formal total synthesis of ganocins A-B, and ganocochlearins C-D, has been achieved. Additionally, a novel synthetic approach for the synthesis of 6,6-disubstituted 6H-dibenzo[b,d]pyran and its analogues has been developed through palladium(II)-catalyzed Wacker-type/cross-coupling cascade reactions.

Transcriptional effects of carbon and nitrogen starvation on Ganoderma boninense, an oil palm phytopathogen.

BACKGROUND: Ganoderma boninense is a phytopathogen of oil palm, causing basal and upper stem rot diseases. METHODS: The genome sequence was used as a reference to study gene expression during growth in a starved carbon (C) and nitrogen (N) environment with minimal sugar and sawdust as initial energy sources. This study was conducted to mimic possible limitations of the C-N nutrient sources during the growth of G. boninense in oil palm plantations. RESULTS: Genome sequencing of an isolate collected from a palm tree in West Malaysia generated an assembly of 67.12 Mb encoding 19,851 predicted genes. Transcriptomic analysis from a time course experiment during growth in this starvation media identified differentially expressed genes (DEGs) that were found to be associated with 29 metabolic pathways. During the active growth phase, 26 DEGs were related to four pathways, including secondary metabolite biosynthesis, carbohydrate metabolism, glycan metabolism and mycotoxin biosynthesis. G. boninense genes involved in the carbohydrate metabolism pathway that contribute to the degradation of plant cell walls were up-regulated. Interestingly, several genes associated with the mycotoxin biosynthesis pathway were identified as playing a possible role in pathogen-host interaction. In addition, metabolomics analysis revealed six metabolites, maltose, xylobiose, glucooligosaccharide, glycylproline, dimethylfumaric acid and arabitol that were up-regulated on Day2 of the time course experiment. CONCLUSIONS: This study provides information on genes expressed by G. boninense in metabolic pathways that may play a role in the initial infection of the host.

Genetic Structure of Ganoderma boninense Populations Associated with Oil Palm at Neighboring Fields with Different Planting Ages in Malaysia.

Ganoderma boninense is a basidiomycete pathogen of African oil palm (Elaeis guineensis) and the causal agent of basal stem rot (BSR) disease, which is the most destructive fungal disease of oil palm in Southeast Asia. The disease is fatal for infected palms and can result in 50 to 80% losses in oil yields because of a reduction in productive life span and a yield decline of infected oil palms. In this study, G. boninense isolates collected from different locations and planting blocks with different palm ages were molecularly characterized using microsatellite genotyping. Results showed high pathogen genetic diversity (He = 0.67 to 0.74) among planting blocks and between oil palm estates. Two nearby planting blocks with similar planting ages (i.e., 1999 and 2001) had a similar percentage of BSR incidence (>20%) but showed distinct Ganoderma genetic structure as detected using STRUCTURE. Similar results were obtained from another trial site where planting blocks differing in planting age but located only less than 1 km apart showed a diverse genetic background. The pathogen genetic admixture of the oldest planting (>30% BSR incidence) differed significantly from the younger planting (1.8 to 2.8% BSR incidence, breeding trial block), suggesting that the host-pathogen genotype interaction may impact the Ganoderma genetic variation over time. The genetic structure of G. boninense, as revealed in this study, implies positive selection resulting from the pathogen genetic variation, host-pathogen interaction, and possible introductions of novel genetic variants (through spores) from adjacent plantings. These findings offer new insights into the genetic changes of G. boninense over time. The information is essential to design disease management strategies and breeding for BSR resistance in oil palm.

Ganonorsterone A, a norsteroid from the medicinal fungus Ganoderma lingzhi.

Phytochemical investigation on the fruiting bodies of the medicinal fungus Ganoderma lingzhi led to the isolation of a new norsteroid, namely ganonorsterone A (1), together with one known steroid, cyathisterol (2). The structure and absolute configuration of compound 1 were assigned by extensive analysis of MS, NMR data, and quantum-chemical calculations including electronic circular dichroism (ECD) and calculated 13C NMR-DP4+ analysis. Bioassay results showed that compound 1 displayed moderate inhibition on NO production in RAW 264.7 macrophages.

Ganoderma lucidum-Derived Meroterpenoids Show Anti-Inflammatory Activity In Vitro.

Ganoderma lucidum, known as the "herb of spiritual potency", is used for the treatment and prevention of various diseases, but the responsible constituents for its therapeutic effects are largely unknown. For the purpose of obtaining insight into the chemical and biological profiling of meroterpenoids in G. lucidum, various chromatographic approaches were utilized for the title fungus. As a result, six undescribed meroterpenoids, chizhienes A-F (1-6), containing two pairs of enantiomers (4 and 5), were isolated. Their structures were identified using spectroscopic and computational methods. In addition, the anti-inflammatory activities of all the isolates were evaluated by Western blot analysis in LPS-induced macrophage cells (RAW264.7), showing that 1 and 3 could dose dependently inhibit iNOS but not COX-2 expression. Further, 1 and 3 were found to inhibit nitric oxide (NO) production using the Greiss reagent test. The current study will aid in enriching the structural and biological diversity of Ganoderma-derived meroterpenoids.

Biphasic Fermentation of Trapa bispinosa Shells by Ganoderma sinense and Characterization of Its Polysaccharides and Alcoholic Extract and Analysis of Their Bioactivity.

BACKGROUND: Trapa bispinosa shells (TBs) and its flesh (TBf) have been recognized for their medicinal properties, including antioxidant, antitumor, and immunomodulatory effects. Despite these benefits, TBs are often discarded as waste material, and their applications remain to be further explored. METHODS: In this study, we optimized the solid-state fermentation process of Ganoderma sinense (GS) with TBs using a response surface experiment methodology to obtain the fermented production with the highest water extract rate and DPPH free radical scavenging activity. We prepared and characterized pre-fermentation purified polysaccharides (P1) and post-fermentation purified polysaccharides (P2). Alcoholic extracts before (AE1) and after (AE2) fermentation were analyzed for active components such as polyphenols and flavonoids using UPLC-QTOF-MS/MS (ultra-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry). Mouse macrophages (RAW 264.7) were employed to compare the immune-stimulating ability of polysaccharides and the antioxidant activity of AE1 and AE2. RESULTS: Optimal fermentation conditions comprised a duration of 2 days, a temperature of 14 °C, and a humidity of 77%. The peak water extract yield and DPPH free radical scavenging rate of the water extract from TBs fermented by GS were observed under these conditions. The enhanced activity may be attributed to changes in the polysaccharide structure and the components of the alcoholic extract. The P2 treatment group indicated more secretion of RAW 264.7 cells of NO, iNOS, IL-2, IL-10, and TNF-α than P1, which shows that the polysaccharides demonstrated increased immune-stimulating ability, with their effect linked to the NF-кB pathway. Moreover, the results of the AE2 treatment group indicated that secretion of RAW 264.7 cells of T-AOC and T-SOD increased and MDA decreased, which shows that the alcoholic extract demonstrated enhanced antioxidant activity, with its effect linked to the Nrf2/Keap1-ARE pathway. CONCLUSIONS: Biphasic fermentation of Trapa bispinosa shells by Ganoderma sinense could change the composition and structure of the polysaccharides and the composition of the alcoholic extract, which could increase the products' immunomodulatory and antioxidant activity.

Use of mushrooms as antioxidants in a lipid oxidation model under indirect and direct oxidation tests: ethanolic extracts of Ganoderma resinaceum and Phlebopus bruchii.

BACKGROUND: Foods contain lipids that are easily susceptible to oxidation, which can modify their sensory properties. Although these compounds provide characteristic flavours and odours, there are also unwanted compounds, such as volatile secondary oxidation products, representing a recurring problem for both the industry and consumers. Synthetic antioxidants are often employed to prevent this but their chronic consumption can be detrimental to human health. The present study evaluates the antioxidant potential of ethanolic extracts from Ganoderma resinaceum and Phlebopus bruchii using an accelerated oxidation test. RESULTS: The composition profile of the extracts was investigated, identifying the presence of tryptophan, quinic acid, caffeic acid and 3,4-dihydroxyphenylglycol-phenolic acid. The antioxidant capacity of the extracts was compared with that of butylated hydroxytoluene (BHT) in sunflower oil that was oven-heated at 60 °C. Chemical (peroxide value, p-anisidine value and conjugated dienes) and volatile (2-octenal, 2-heptenal and 2,4-decadienal) indicators were measured over 28 days. The peroxide value decreased for both extracts at a similar level to that of BHT 0.02% w/w, and conjugate dienes decreased in the presence of G. resinaceum 0.1% w/w. Meanwhile, p-anisidine exhibited a slightly greater decrease for P. bruchii 0.1% w/w than for BHT. The sample with 0.1% w/w of extracts showed a reduction in volatile secondary oxidation compounds, indicating significant antioxidant activity. CONCLUSION: Based on these results, both extracts could be proposed as potential antioxidants in foods with a high lipid content. © 2024 Society of Chemical Industry.

Application of sequential design for enhanced L-asparaginase synthesis from Ganoderma australe GPC191.

With an increasing demand for L-asparaginase in pharmaceutical and food sectors for its cytostatic and acrylamide-reducing qualities, there's a need to discover novel, highly productive enzyme sources with improved pharmacokinetic profiles. Keeping this in mind, the present study aimed at maximizing the potential of Ganoderma australe GPC191 to produce L-asparaginase by fermentation medium optimization using statistical validation. Of the 11 physicochemical parameters evaluated under submerged fermentation conditions through one-factor-at-a-time approach and Plackett-Burman design, only four parameters (inoculum load, L-asparagine, soybean meal, and initial pH) influenced L-asparaginase production, significantly (p < 0.001). The optimal levels and interaction effects of these on the overall production were further evaluated by the central composite rotatable design of response surface methodology. Post-optimization, 27.34 U/mL was predicted as the maximum activity at pH 7 with 5n inoculum load and 15 g/L each of L-asparagine and soybean meal. Experimental validation yielded an activity of 28.52 U/mL, indicating an overall 18.17-fold increase from the unoptimized stage. To our knowledge, this is the first report signifying the L-asparaginase production aptitude of G. australe with sequential statistical validation using agricultural waste, which can serve as a model to enhance its yields, offering a sustainable and cost-effective solution for industrial application.

Identification a novel Ganoderma FIP gene from Ganoderma capense and its functional expression in Pichia pastoris.

Ganoderma capense is a precious medicinal fungus in China. In this study, a novel fungal immunomodulatory protein gene, named as FIP-gca, was cloned from G. capense by homologous cloning. Sequencing analysis indicated that FIP-gca was composed of 336 bp, which encoded a polypeptide of 110 amino acids. Protein sequence blasting and phylogenetic analysis showed that FIP-gca shared homology with other Ganoderma FIPs. FIP-gca was effectively expressed in Pichia pastoris GS115 at an expression level of 166.8 mg/L and purified using HisTrap™ fast-flow prepack columns. The immunomodulation capacity of rFIP-gca was demonstrated by that rFIP-gca could obviously stimulate cell proliferation and increase IL-2 secretion of murine spleen lymphocytes. Besides, antitumor activity of rFIP-gca towards human stomach cancer AGS cell line was evaluated in vitro. Cell wound scratch assay proved that rFIP-gca could inhibit migration of AGS cells. And flow cytometry assay revealed that rFIP-gca could significantly induce apoptosis of AGS cells. rFIP-gca was able to induce 18.12% and 22.29% cell apoptosis at 0.3 µM and 0.6 µM, respectively. Conclusively, the novel FIP-gca gene from G. capense has been functionally expressed in Pichia and rFIP-gca exhibited ideal immunomodulation and anti-tumour activities, which implies its potential application and study in future.

Bidirectional Solid-State Fermentation of Highland Barley by Edible Fungi to Improve Its Functional Components, Antioxidant Activity and Texture Characteristics.

In food industry, the characteristics of food substrate could be improved through its bidirectional solid-state fermentation (BSF) by fungi, because the functional components were produced during BSF. Six edible fungi were selected for BSF to study their effects on highland barley properties, such as functional components, antioxidant activity, and texture characteristics. After BSF, the triterpenes content in Ganoderma lucidum and Ganoderma leucocontextum samples increased by 76.57 and 205.98%, respectively, and the flavonoids content increased by 62.40% (Phellinus igniarius). Protein content in all tests increased significantly, with a maximal increase of 406.11% (P. igniarius). Proportion of indispensable amino acids increased significantly, with the maximum increase of 28.22%. Lysine content increased largest by 437.34% to 3.310 mg/g (Flammulina velutipes). For antioxidant activity, ABTS radical scavenging activity showed the maximal improvement, with an increase of 1268.95%. Low-field NMR results indicated a changed water status of highland barley after fermentation, which could result in changes in texture characteristics of highland barley. Texture analysis showed that the hardness and chewiness of the fermented product decreased markedly especially in Ganoderma lucidum sample with a decrease of 77.96% and 58.60%, respectively. The decrease indicated a significant improvement in the taste of highland barley. The results showed that BSF is an effective technology to increase the quality of highland barley and provide a new direction for the production of functional foods.

Structural diversity, hypothetical biosynthesis, chemical synthesis, and biological activity of Ganoderma meroterpenoids.

Covering: 2018 to 2022Meroterpenoids found in fungal species of the genus Ganoderma and known as Ganoderma meroterpenoids (GMs) are substances composed of a 1,2,4-trisubstituted benzene and a polyunsaturated side chain. These substances have attracted the attention of chemists and pharmacologists due to their diverse structures and significant bioactivity. In this review, we present the structures and possible biosynthesis of representative GMs newly found from 2018 to 2022, as well as chemical synthesis and biological activity of some interesting GMs. We propose for the first time a plausible biosynthetic pathway for GMs, which will certainly motivate further research on the biosynthetic pathway in Ganoderma species, as well as on chemical synthesis of GMs as important bioactive compounds for the purpose of drug development.

Nondestructive and Quantitative Analysis of Cell Wall Regeneration in the Medicinal Macrofungus Ganoderma lingzhi by a Membrane-Fusing Fluorescent Probe.

Ganoderma is a prize medicinal macrofungus with a broad range of pharmaceutical values. To date, various attempts have been made to cultivate Ganoderma to improve the production of secondary metabolites with pharmacological activity. Among the adopted techniques, protoplast preparation and regeneration are indispensable. However, the evaluation of protoplasts and regenerated cell walls usually relies on electron microscopy assays, which require time-consuming and destructive sample preparation and merely provide localized information in the selected area. In contrast, fluorescence assays enable sensitive real-time detection and imaging in vivo. They can also be applied to flow cytometry, providing a collective overview of every cell in a sample. However, for macrofungi such as Ganoderma, the fluorescence analysis of protoplasts and regenerated cell walls is difficult owing to the hindrance of the homologous fluorescent protein expression and the lack of an appropriate fluorescence marker. Herein, a specific plasma membrane probe, TAMRA perfluorocarbon nucleic acid probe (TPFN), is proposed for the nondestructive and quantitative fluorescence analysis of cell wall regeneration. Exploiting the perfluorocarbon membrane-anchoring chains, hydrophilic nucleic acid linker, and fluorescent dye TAMRA, the probe is proven to be selective, soluble, and stable, enabling rapid fluorescence detection of a protoplast sample free of transgenic expression or immune staining. Based on the TPFN and flow cytometry techniques, a quantitative approach is constructed to monitor the process of cell wall growth in a fast, quantitative, and high-throughout manner, and the obtained results are consistent with those of conventional electron microscopy. In principle, with slight modifications or integration, the proposed probe and approach can be adapted to the preparation of cell protoplasts, inspection of cell wall integrity under environmental stress, and programmable membrane engineering for cytobiology and physiology research.

Comparative proteomic analysis of two divergent strains provides insights into thermotolerance mechanisms of Ganoderma lingzhi.

Heat stress (HS) is a major abiotic factor influencing fungal growth and metabolism. However, the genetic basis of thermotolerance in Ganoderma lingzhi (G. lingzhi) remains largely unknown. In this study, we investigated the thermotolerance capacities of 21 G. lingzhi strains and screened the thermo-tolerant (S566) and heat-sensitive (Z381) strains. The mycelia of S566 and Z381 were collected and subjected to a tandem mass tag (TMT)-based proteome assay. We identified 1493 differentially expressed proteins (DEPs), with 376 and 395 DEPs specific to the heat-tolerant and heat-susceptible genotypes, respectively. In the heat-tolerant genotype, upregulated proteins were linked to stimulus regulation and response. Proteins related to oxidative phosphorylation, glycosylphosphatidylinositol-anchor biosynthesis, and cell wall macromolecule metabolism were downregulated in susceptible genotypes. After HS, the mycelial growth of the heat-sensitive Z381 strain was inhibited, and mitochondrial cristae and cell wall integrity of this strain were severely impaired, suggesting that HS may inhibit mycelial growth of Z381 by damaging the cell wall and mitochondrial structure. Furthermore, thermotolerance-related regulatory pathways were explored by analyzing the protein-protein interaction network of DEPs considered to participate in the controlling the thermotolerance capacity. This study provides insights into G. lingzhi thermotolerance mechanisms and a basis for breeding a thermotolerant germplasm bank for G. lingzhi and other fungi.

Agitation role (Dissolved Oxygen) in production of laccase from newly identified Ganoderma multistipitatum sp. nov. and its effect on mycelium morphology.

BACKGROUND: Agitation speed influenced the production rate of laccase. Orbital speed not only influenced the enzyme production, but was also effective to dissolve the oxygen during growth of mycelium, spores, and chlamydospores. Shear effects of speed greatly influenced the morphology of mycelium. METHODS: Ganoderma multistipitatum was identified by ITS marker. Phylogenetic tree was constructed for species identification. Qualitatively by plate method contained guaiacol indicator, while quantitatively by submerged fermentation and Central Composite Design applied on agitation parameter for maximum laccase potential of this species. The effects of agitation speed on mycelium morphology were observed under compound and scanning electron microscope. RESULTS: Statistical optimization of agitation conditions were performed by using response surface methodology to enhance the production of laccase from Ganoderma multistipitatum sp. nov. Maximum laccase yield (19.44 × 105 ± 0.28 U/L) was obtained at 150 rpm grown culture, which was higher than predicted value of laccase production (19.18 × 105 U/L) under aerobic conditions (150 rpm). The 150 rpm provided the continuous flush of oxygen. The DO (dissolved oxygen) was maximum (65%) for "27 h" incubation at 150 rpm during laccase synthesis. The statistical value of laccase production was minimum under anaerobic or nearly static condition of 50 rpm. The predicted (12.78 × 105 U/L) and obtained (12.82 × 105 U/L) yield was low at 50 rpm. Optimization of orbital shaking for aeration conditions were performed by the use of "Response Surface Methodology". The submerged shaking flasks were utilized as a nutrients growth medium to maximize the production of laccase from G. multistipitatum. The minimum incubation time highly influenced the laccase yield from 7 to 15 days via utilization of less cost-effective medium under a promising and eco-friendly method. The morphological effects of rpm on mycelium were examined under compound and scanning electron microscopy. Higher rpm (200, 230) shear the mycelium, while 150 to 200 rpm exhibited smoother and highly dense branches of mycelia. CONCLUSION: The shear forces of 200 rpm caused the damages of mycelium and cells autolysis with less laccase production. This study concluded that 150 rpm saved the life of mycelium and enhanced the production rate of enzymes.

Induced expression of Ganoderma boninense Lanosterol 14α-Demethylase (ERG11) during interaction with oil palm.

BACKGROUND: The basidiomycete fungus, Ganoderma boninense is the main contributor to oil palm Basal Stem Rot (BSR) in Malaysia and Indonesia. Lanosterol 14α-Demethylase (ERG11) is a key enzyme involved in biosynthesis of ergosterol, which is an important component in the fungal cell membrane. The Azole group fungicides are effective against pathogenic fungi including G. boninense by inhibiting the ERG11 activity. However, the work on molecular characterization of G. boninense ERG11 is still unavailable today. METHODS AND RESULTS: This study aimed to isolate and characterize the full-length cDNA encoding ERG11 from G. boninense. The G. boninense ERG11 gene expression during interaction with oil palm was also studied. A full-length 1860 bp cDNA encoding ERG11 was successfully isolated from G. boninense. The G. boninense ERG11 shared 91% similarity to ERG11 from other basidiomycete fungi. The protein structure homology modeling of GbERG11 was analyzed using the SWISS-MODEL workspace. Southern blot and genome data analyses showed that there is only a single copy of ERG11 gene in the G. boninense genome. Based on the in-vitro inoculation study, the ERG11 gene expression in G. boninense has shown almost 2-fold upregulation with the presence of oil palm. CONCLUSION: This study provided molecular information and characterization study on the G. boninense ERG11 and this knowledge could be used to design effective control measures to tackle the BSR disease of oil palm.

Antimalarial Lanostane Dimers from Artificially Cultivated Fruiting Bodies of Ganoderma weberianum.

Investigation of cultivated fruiting bodies of Ganoderma weberianum led to the isolation of 11 previously unreported lanostane dimers, ganoweberianones C (3a), D (4a), E (5a), F (6a), G (7a), and H (8a) and isoganoweberianones A (1b), B (2b), D (4b), G (7b), and H (8b). Six new ganodermanontriol derivatives as three pairs of diastereomers (11/12, 13/14, and 15/16) and five new ganoweberianic acids (17-21) were also isolated. A method for semisynthesis of lanostane dimers by condensation of natural lanostanes was established, which was utilized in the structure elucidation and NMR data assignments of the undescribed natural lanostane dimers. Ganoweberianone D (4a) and isoganoweberianone D (4b) showed significant antimalarial activity against Plasmodium falciparum K1 (multidrug-resistant strain) with IC50 values of 0.057 and 0.035 µM, respectively, whereas their cytotoxicity to Vero cells was weaker (IC50 8.1 and 19 µM, respectively).