Heat Stress Modulates Mycelium Growth, Heat Shock Protein Expression, Ganoderic Acid Biosynthesis, and Hyphal Branching of Ganoderma lucidum via Cytosolic Ca2.

UNLABELLED: Heat stress (HS) influences the growth and development of organisms. Thus, a comprehensive understanding of how organisms sense HS and respond to it is required. Ganoderma lucidum, a higher basidiomycete with bioactive secondary metabolites, has become a potential model system due to the complete sequencing of its genome, transgenic systems, and reliable reverse genetic tools. In this study, we found that HS inhibited mycelium growth, reduced hyphal branching, and induced the accumulation of ganoderic acid biosynthesis and heat shock proteins (HSPs) in G. lucidum Our data showed that HS induced a significant increase in cytosolic Ca(2+) concentration. Further evidence showed that Ca(2+) might be a factor in the HS-mediated regulation of hyphal branching, ganoderic acid (GA) biosynthesis, and the accumulation of HSPs. Our results further showed that the calcium-permeable channel gene (cch)-silenced and phosphoinositide-specific phospholipase gene (plc)-silenced strains reduced the HS-induced increase in HSP expression compared with that observed for the wild type (WT). This study demonstrates that cytosolic Ca(2+) participates in heat shock signal transduction and regulates downstream events in filamentous fungi. IMPORTANCE: Ganoderma lucidum, a higher basidiomycete with bioactive secondary metabolites, has become a potential model system for evaluating how environmental factors regulate the development and secondary metabolism of basidiomycetes. Heat stress (HS) is an important environmental challenge. In this study, we found that HS inhibited mycelium growth, reduced hyphal branching, and induced HSP expression and ganoderic acid biosynthesis in G. lucidum Further evidence showed that Ca(2+) might be a factor in the HS-mediated regulation of hyphal branching, GA biosynthesis, and the accumulation of HSPs. This study demonstrates that cytosolic Ca(2+) participates in heat shock signal transduction and regulates downstream events in filamentous fungi. Our research offers a new way to understand the mechanism underlying the physiological and metabolic responses to other environmental factors in G. lucidum This research may also provide the basis for heat shock signal transduction studies of other fungi.

[Effect of different light of LED light quality on growth and antioxidant enzyme activities of Ganoderma lucidum].

OBJECTIVE: To study the effect of light quality on growth, antioxidant enzyme activities of Ganoderma lucidum mycelium. METHOD: G. lucidum mycelium was cultured under different light qualities by light emitting diodes (LED). The growth G. lucidum mycelium was observed and antioxidant enzyme activities was determined in different growth periods. RESULT: Under the red LED, the blue LED and dark condition (CK), the mycelium grew faster than that under other light qualities. The white LED resulted in a largest increase in the amount of the mycelium and always kept the activities of CAT high level. Major fluctuations of POD activities emerged under the green LED, while enhanced severely in the late phase. Under the yellow LED, the activities of SOD appeared high level. However, SOD activities on dark (CK) raised obviously in late period. At the late stage, the content of mycelium polysaccharides was significant higher than that under the blue LED. CONCLUSION: The light quality could influence the growth and metabolism of G. lucidum mycelium.

[Effect of light quality on growth and polysaccharides content of Ganoderma lucidum].

OBJECTIVE: To study the effect of light quality on growth, polysaccharides content of Ganoderma lucidum. METHOD: G. lucidum was planted under different light qualities. The growth of G. lucidum was observed and polysaccharides content was determined in different growth periods. RESULT: There were significant differences in the form of the thickness and the numbers of ringed of the fruit before the ejection. The maximum content of G. lucidum polysaccharides under different light qualities were appeared in different periods. The content of polysaccharides was higher than that of CK under blue light in the period of budding, parachute phase, after spore ejection. The content of polysaccharides was increased under green light in the period of growth stage. The yield of fruits and spores were different in different light qualities. CONCLUSION: The light quality could increase the content of polysaccharides in G. lucidum.

Conditions Affecting Lingzhi or Reishi Medicinal Mushroom Ganoderma lucidum (Agaricomycetes) Basidiome Quality, Morphogenesis, and Biodegradation of Wood By-products in Argentina.

Solid-state fermentation (SSF) with the medicinal higher Basidiomycete Ganoderma lucidum was studied as a strategy to use pine (Pinus radiata D. Don) and poplar (Populus nigra L.) wood chips and sawdust. Fruiting bodies were produced and the value of the biotransformed substrate was assessed. The highest mushroom yield (63 g dry weight per kilogram of dry substrate) was obtained with poplar sawdust and wood chips. Immersion of the bioreactors was a simple watering method that obtained suitable yields. Two morphological types were induced using 2 different incandescent light intensities. High light irradiation induced the highest valued mushroom morphology (as a whole product). Time course study of substrate biodegradation and mycelial growth dynamics indicated that the trophophase lasted 20 days and presented laccase activity of 0.01-0.03 units · g-1. The activity at idiophase was 10 times higher. Aqueous and alkali extracts, as well as carbohydrase enzyme profile activity, revealed differences in the properties of the residual substrate; some related to the substrate source are considered to be of concern for further use of this pretreated biomass. In view of the results obtained, we propose use of SSF of pine and poplar with G. lucidum to profitably recycle softwood by-products from the timber industry.

Development of antler-type fruiting bodies of Ganoderma lucidum and determination of its biochemical properties.

Following the importance of antler-type fruiting bodies of Ganoderma lucidum, in this study, the impact of main growth parameters such as ventilation and light on the development of antler-type fruiting bodies has been investigated together with the determination of physico-chemical properties of antler fruiting bodies. For this, the primordia bags of G. lucidum were kept under controlled ventilation to adjust the CO2 produced by the mushrooms owing to its respiration under light and dark conditions. The bioactive compounds such as phenolics, flavonoids, water-soluble polysaccharides and ganoderic acid showed a two-fold increase in the antler-type fruiting bodies as compared to normal kidney-shaped fruiting bodies. It is assumed from this study that the antler type fruiting bodies are developed due to restricted ventilation which causes an increase in the level of CO2 gas in the air as a result of respiration of mushroom. The shape and colour of antler fruiting bodies again dependent on the light provided in the growth chamber. This study also proves that with the manipulation of light and ventilation antler-type fruiting bodies of G. lucidum could be developed with higher quantity of bioactive compounds and with higher antioxidant potential.

Identification of Reference Genes and Analysis of Heat Shock Protein Gene Expression in Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum, after Exposure to Heat Stress.

Ganoderma lucidum has been considered an emerging model species for studying how environmental factors regulate the growth, development, and secondary metabolism of Basidiomycetes. Heat stress, which is one of the most important environmental abiotic stresses, seriously affects the growth, development, and yield of microorganisms. Understanding the response to heat stress has gradually become a hotspot in microorganism research. But suitable reference genes for expression analysis under heat stress have not been reported in G. lucidum. In this study, we systematically identified 11 candidate reference genes that were measured using reverse transcriptase quantitative polymerase chain reaction, and the gene expression stability was analyzed under heat stress conditions using geNorm and NormFinder. The results show that 5 reference genes-CYP and TIF, followed by UCE2, ACTIN, and UBQ1-are the most stable genes under our experimental conditions. Moreover, the relative expression levels of 3 heat stress response genes (hsp17.4, hsp70, and hsp90) were analyzed under heat stress conditions with different normalization strategies. The results show that use of a gene with unstable expression (SAND) as the reference gene leads to biased data and misinterpretations of the target gene expression level under heat stress.

A novel three-stage light irradiation strategy in the submerged fermentation of medicinal mushroom Ganoderma lucidum for the efficient production of ganoderic acid and Ganoderma polysaccharides.

A novel three-stage light irradiation strategy in the submerged fermentation of medicinal mushroom Ganoderma lucidum for the efficient production of bioactive metabolites ganoderic acid (GA) and Ganoderma polysaccharides was developed. Significance of light quality, i.e., blue light (390-500 nm, lambda(max) = 470 nm), red light (560-700 nm, lambda(max) = 625 nm), and white light (400-740 nm, lambda(max) = 550 nm), was studied at first. Interestingly, there was a gradual decrease trend of GA content after the culture of day 2 when the maximal GA content was obtained, while GA content decreased slowly under white light irradiation after day 6. The dark environment was favorable to the specific GA biosynthesis (i.e., GA content) before day 6, and after that the optimum was white light irradiation. A relatively lower irradiation density of white light (i.e., 0.94 and 2.82 W/m(2)) was beneficial for the specific GA biosynthesis before day 6, while GA content was higher under higher irradiation density of white light (i.e., 4.70 and 9.40 W/m(2)) at the later-stage of cultivation. 4.70 W/m(2) white light irradiation culture was the best from the viewpoint of GA accumulation. Therefore, a two-stage light irradiation strategy by combing the first 2 days dark culture with the following 4.70 W/m(2) white light irradiation culture was developed. The highest GA production in the two-stage culture was 276.0 +/- 12.5 mg/L, which was increased by 19% compared to 4.70 W/m(2) white light irradiation culture (i.e., 232.4 +/- 15.8 mg/L) and by 178% compared to the dark culture (i.e., 99.4 +/- 1.0 mg/L). Although there still existed a gradual decrease trend of GA content after day 2 when the maximal GA content was obtained in the two-stage culture. Following three-stage light irradiation strategy was further demonstrated in order to turn around the sharp decrease of GA content after day 2. The first-stage was the 2-day dark culture; the second-stage was the following six-day 0.94 W/m(2) white light irradiation culture, and the third-stage was 4.70 W/m(2) white light irradiation culture until the end of fermentation. During the three-stage culture of G. lucidum, the gradual decrease trend of GA content after day 2 was turned around, which suggested that 0.94 W/m(2) white light irradiation was beneficial for the metabolic flux towards the GA biosynthesis. The maximal GA content of 3.1 +/- 0.1 mg/100 mg DW was obtained, which was higher by 41% compared to the two-stage culture. The maximal GA production (i.e., 466.3 +/- 24.1 mg/L) and productivity (i.e., 38.9 mg/L per day) in the three-stage culture were 69 and 101% higher than those obtained in the two-stage culture. This is the first report investigating the significance of light irradiation on the medicinal mushroom submerged fermentation. Such work is very helpful to other mushroom fermentations for useful metabolite production.