Transcriptome Reveals Roles of Lignin-Modifying Enzymes and Abscisic Acid in the Symbiosis of Mycena and Gastrodia elata.

Gastrodia elata is a well-known medicinal and heterotrophic orchid. Its germination, limited by the impermeability of seed coat lignin and inhibition by abscisic acid (ABA), is triggered by symbiosis with fungi such as Mycena spp. However, the molecular mechanisms of lignin degradation by Mycena and ABA biosynthesis and signaling in G. elata remain unclear. In order to gain insights into these two processes, this study analyzed the transcriptomes of these organisms during their dynamic symbiosis. Among the 25 lignin-modifying enzyme genes in Mycena, two ligninolytic class II peroxidases and two laccases were significantly upregulated, most likely enabling Mycena hyphae to break through the lignin seed coats of G. elata. Genes related to reduced virulence and loss of pathogenicity in Mycena accounted for more than half of annotated genes, presumably contributing to symbiosis. After coculture, upregulated genes outnumbered downregulated genes in G. elata seeds, suggesting slightly increased biological activity, while Mycena hyphae had fewer upregulated than downregulated genes, indicating decreased biological activity. ABA biosynthesis in G. elata was reduced by the downregulated expression of 9-cis-epoxycarotenoid dioxygenase (NCED-2), and ABA signaling was blocked by the downregulated expression of a receptor protein (PYL12-like). This is the first report to describe the role of NCED-2 and PYL12-like in breaking G. elata seed dormancy by reducing the synthesis and blocking the signaling of the germination inhibitor ABA. This study provides a theoretical basis for screening germination fungi to identify effective symbionts and for reducing ABA inhibition of G. elata seed germination.

[Ecological benefits of Gastrodia elata-Phallus impudicus sequential planting pattern].

Gastrodia elata is a kind of precious traditional Chinese medicine. In the process of cultivation of G. elata, due to the influence of continuous cropping obstacles and other factors, the fungus materials and land that have been planted with G. elata are often abandoned, resulting in a great waste of resources. Based on the planting characteristics of G. elata and Phallus impudicus and the previous research experience in ecological agriculture, this paper analyzed the ecological adaptability characteristics of G. elata and P. impudicus, and summarized the key techniques of the G. elata-P. impudicus sequential planting pattern. Keeping track of the planting area, fungus-growing materials consumption and market sales of G. elata-P. impudicus sequential planting pattern, the ecological benefits of G. elata-P. impudicus sequential planting pattern from the aspects of utilization rate of fungus-growing materials were analyzed, the value of land resources per unit area, ecological environmental protection, labor cost and economic benefits were consi-dered. The technical principle of G. elata-P. impudicus sequential planting pattern was expounded according to their ecological habit, the season of harvest and planting, the difference of composition of fungus-growing materials, and the microbial ecology. The sequential planting pattern of G. elata-P. impudicus not only realized the double production of medicinal materials and edible fungi, reduced the waste of old fungus-growing materials, but also transformed the energy from nutrition-supplied fungi to edible and medicinal fungi, which guaranteed the ecological recycling and utilization of G. elata in the process of cultivation.

[Effect of Gastrodiae elata-Phallus impudicus sequential planting pattern on soil microbial community structure].

Gastrodia elata is a heterotrophic plant that needed to be symbiotic with Armillaria. The obstacle of continuous cropping in G. elata is serious during the G. elata cultivation, and the mechanism of obstacle in G. elata continuous cropping had not been solved. The planting of G. elata-Phallus impudicus is a new sequential planting pattern adopted in Guizhou province, but the effect of the cultivation on soil microbial community structure is still unclear. In this study, we collected four soil samples for the research including the soil without planted G. elata as control(CK), rhizosphere soil samples tightly adhering to the G. elata surface(GE), rhizosphere soil samples tightly adhering to Armillaria which was symbiotic with G. elata(AGE), the rhizosphere soil of P. impudicus planting after G. elata cultivation(PI). In order to explore the mechanism, the research study on the soil of G. elata-P. impudicus by using ITS and 16 S rDNA high-throughput sequencing technologies to detect soil microbial community structure including fungi and bacteria in the soil of CK, AGE, GE and PI. OTU clustering and PCA analysis of soil samples showed that the soil microbial diversity was relatively similar in AGE and GE. And the soil microbial in PI and CK clustered together. The results showed that AGE and GE had similar soil microbial diversity, as well as PI and CK. Compared with CK, the soil microbial diversity and abundance in AGE and GE were significantly increased. But the microbial diversity and abundance decreased in PI compared with AGE and GE. The annotation indicated that the abundance of Basidiomycota, Acidobacteria and Chloroflexi decreased, and that of Ascomycota, Zygomycota and Proteobacteria increased in AGE and GE compared with CK. In contrast to AGE and GE, PI was the opposite. The abundance of Basidiomycota, Acidobacteria and Chloroflexi increased in PI compared with AGE and GE. The abundance of microorganisms in the soil of PI and CK was similar. In addition, the co-culture of Armillaria and P. impudicus indicated that P. impudicus had obvious antagonistic effects on the growth of Armillaria. Therefore, it is speculated that the mechanism of G. elata-P. impudicus planting pattern related to the change of soil microbial. And we supposed that P. impudicus might inhibit the growth of Armillaria and change the soil microbial community structure and the abundance of soil microbial. And the soil microbial community structure was restored to a state close to that of uncultivated G. elata. Thus, the structure of soil microbial community planting G. elata could be restored by P. impudicus planting.

[Dissection of seed development of Gastrodia elata at different temperatures].

The study is aimed to create seed materials and dissect the molecular mechanism of sexual propagation of Gastrodia elata. In this research, thirteen characteristics of flowers, flower stem, fruits, seeds and embryo of G.elata f. glauca and G.elata f. elata after bolting at room temperature(RT) and constant temperature(CT, 22 ℃) were determined. It was found that the constant temperature condition could prolong the bolting duration of G.elata and increased the number of flowers, while the variety of G.elata only affected the bolting duration, but had no effect on the number of flowers, and the G.elata f. elata was more likely to bolting than the G.elata f. glauca. The variety of G.elata was the main factor affecting the time of dehiscent fruit of G.elata, the temperature was the main factor affecting the fruits number and fruits diameter, and the constant temperature was more conducive to the fruits shape of G.elata than the room temperature. There was no significant difference in seed phenotype of G.elata varieties, but the seed embryo of G.elata seeds cultivated at constant temperature was fuller than that of G.elata cultivated at room temperature, and temperature had less influence on the seed phenotype of G.elata. But it was interesting to find that temperature and varieties had greater influence on the seed embryo of G.elata, constant temperature cultivation was more conducive to the formation of seed embryo of G.elata, and more the seed embryo of G.elata f. elata was easier to form than the seed embryo of G.elata f. glauca. However, the development of seeds and embryos of G.elata was significantly affected, and the development of seeds and embryos of G.elata f. glauca was more sensitive to temperature than G.elata f. elata. The research suggested that it is advisable for G.elata to produce seed materials by bolting at constant temperature(22 ℃).

Dissection of seed development of Gastrodia elata at different temperatures / 中国中药杂志

The study is aimed to create seed materials and dissect the molecular mechanism of sexual propagation of Gastrodia elata. In this research, thirteen characteristics of flowers, flower stem, fruits, seeds and embryo of G.elata f. glauca and G.elata f. elata after bolting at room temperature(RT) and constant temperature(CT, 22 ℃) were determined. It was found that the constant temperature condition could prolong the bolting duration of G.elata and increased the number of flowers, while the variety of G.elata only affected the bolting duration, but had no effect on the number of flowers, and the G.elata f. elata was more likely to bolting than the G.elata f. glauca. The variety of G.elata was the main factor affecting the time of dehiscent fruit of G.elata, the temperature was the main factor affecting the fruits number and fruits diameter, and the constant temperature was more conducive to the fruits shape of G.elata than the room temperature. There was no significant difference in seed phenotype of G.elata varieties, but the seed embryo of G.elata seeds cultivated at constant temperature was fuller than that of G.elata cultivated at room temperature, and temperature had less influence on the seed phenotype of G.elata. But it was interesting to find that temperature and varieties had greater influence on the seed embryo of G.elata, constant temperature cultivation was more conducive to the formation of seed embryo of G.elata, and more the seed embryo of G.elata f. elata was easier to form than the seed embryo of G.elata f. glauca. However, the development of seeds and embryos of G.elata was significantly affected, and the development of seeds and embryos of G.elata f. glauca was more sensitive to temperature than G.elata f. elata. The research suggested that it is advisable for G.elata to produce seed materials by bolting at constant temperature(22 ℃).

Ecological benefits of Gastrodia elata-Phallus impudicus sequential planting pattern / 中国中药杂志

Gastrodia elata is a kind of precious traditional Chinese medicine. In the process of cultivation of G. elata, due to the influence of continuous cropping obstacles and other factors, the fungus materials and land that have been planted with G. elata are often abandoned, resulting in a great waste of resources. Based on the planting characteristics of G. elata and Phallus impudicus and the previous research experience in ecological agriculture, this paper analyzed the ecological adaptability characteristics of G. elata and P. impudicus, and summarized the key techniques of the G. elata-P. impudicus sequential planting pattern. Keeping track of the planting area, fungus-growing materials consumption and market sales of G. elata-P. impudicus sequential planting pattern, the ecological benefits of G. elata-P. impudicus sequential planting pattern from the aspects of utilization rate of fungus-growing materials were analyzed, the value of land resources per unit area, ecological environmental protection, labor cost and economic benefits were consi-dered. The technical principle of G. elata-P. impudicus sequential planting pattern was expounded according to their ecological habit, the season of harvest and planting, the difference of composition of fungus-growing materials, and the microbial ecology. The sequential planting pattern of G. elata-P. impudicus not only realized the double production of medicinal materials and edible fungi, reduced the waste of old fungus-growing materials, but also transformed the energy from nutrition-supplied fungi to edible and medicinal fungi, which guaranteed the ecological recycling and utilization of G. elata in the process of cultivation.

Dynamics of fungal communities during Gastrodia elata growth.

BACKGROUND: Gastrodia elata is a widely distributed achlorophyllous orchid and is highly valued as both medicine and food. Gastrodia elata produces dust-like seeds and relies on mycorrhizal fungi for its germination and growth. In its life cycle, G. elata is considered to switch from a specific single-fungus relationship (Mycena) to another single-fungus relationship (Armillaria). However, no studies have investigated the changes in the plant-fungus relationship during the growth of G. elata in the wild. In this study, high-throughput sequencing was used to characterize the fungal community of tubers in different growth phases as well as the soils surrounding G. elata. RESULTS: The predominant fungi were Basidiomycota (60.44%) and Ascomycota (26.40%), which exhibited changes in abundance and diversity with the growth phases of G. elata. Diverse basidiomycetes in protocorms (phase P) were Hyphodontia, Sistotrema, Tricholoma, Mingxiaea, Russula, and Mycena, but the community changed from a large proportion of Resinicium bicolor (40%) in rice-like tubers (phase M) to an unidentified Agaricales operational taxonomic unit 1(OTU1,98.45%) in propagation vegetation tubers (phase B). The soil fungi primarily included Simocybe, Psathyrella, Conocybe, and Subulicystidium. Three Mycena OTUs obtained in this study were differentially distributed among the growth phases of G. elata, accounting for less than 1.0% of the total reads, and were phylogenetically close to Mycena epipterygia and M. alexandri. CONCLUSIONS: Our data indicated that G. elata interacts with a broad range of fungi beyond the Mycena genus. These fungi changed with the growth phases of G. elata. In addition, these data suggested that the development of the fungal community during the growth of G. elata was more complex than previously assumed and that at least two different fungi could be involved in development before the arrival of Armillaria.

[Comparative and path analysis between underground tuber yield and agronomic traits in cultivation provenances of Gastrodia elata f. glauca from Zhaotong].

To provide guidance for quality control and variety breeding of Gastrodia elata f. glauca cultivated form in Zhaotong, nine agronomic traits of G. elata f. glauca cultivation form were measured and the traits were analyzed using multiple comparison,correlation analysis, multiple stepwise regression analysis and path analysis. The results of multiple comparison and analysis showed that the number of dot ring and the length-width ratio of tuber were no different in G. elata f. glauca cultivation form. The coefficient of variation about the single tuber fresh weigh, ring spacing and belly button diameter was about 20%. The coefficient of variation about the length of the tuber, the width of the tuber, the thickness of the tuber, the length-width ratio of tuber and the number of dot ring was near 10%. The correlation analysis results showed that the single tuber fresh weigh and the length,the width and the thickness of the tuber were extremely significant positive correlated. The results of multiple stepwise regression analysis and path analysis showed that the length, the width and the thickness of the tuber were the main factors affecting the fresh weight of single tuber,which were the best agronomic traits on high yield of G. elata f. glauca. The length of the tuber,the width of the tuber and the thickness of the tuber, which would be used as indicators of the characteristics of high yield varieties in Zhaotong. The number of dot ring and the length-width ratio of tuber, which would be selected as quality control indicators of the genuine Chinese medicinal materials in Zhaotong.

[Ecological suitability regionalization for Gastrodia elata in Zhaotong based on Maxent and ArcGIS].

In this paper, the potential distribution information and ecological suitability regionalization for Gastrodia elata in Zhaotong were studied based on the climate, terrain, soil and vegetation factors analysis by Maxent and ArcGIS. The results showed that the highly potential distribution (suitability index>0.6) mainly located in Zhaotong, Yunnan province（Zhenxiong,Yiliang and Daguan county, with an area of 2 872 km²ï¼‰, and Bijie, Guizhou province （Hezhang,Bijie,Weining county, 1 251 km²ï¼‰. The AUC of ROC curve was above 0.99, indicating that the predictive results with the Maxent model were highly precise. The main ecological factors determining the potential distribution were the altitude, average rainfall in November, average rainfall in October, vegetation types, average rainfall in March, average rainfall in April,soil types,isothermal characteristic and average rainfall in June. The environmental variables in the highly potential areas were determined as altitude around 1 450-2 200 m,annual average temperature around 18.0-20.4 ℃,annual average precipitation around 900 mm,yellow soil or yellow brown soil,and acid sandy loam or slightly acidic sandy loam.The results will provide valuable references for plantation regionalization and the siting for imitation wild planting of G. elata in Zhaotong.

[Effects of Different Planting Direction and Layer Combination on Gastrodia elata f. elata in Bionic Wild Cultivation].

OBJECTIVE: Combination of different planting direction and layer were set to choose the best technology of cultivation of Gastrodia elata f. elata. METHODS: To improve the yield and quality of Gastrodia elata f. elata, randomized block design experiments were carried out to investigate the yield and quality, and to analyze their economic effectiveness in bionic wild cultivation. RESULTS: Length, width, thickness and weight of southern direction's Gastrodia elata f. elata developed better than the northeast direction. The three planting layer levels on growth effect of Gastrodia elata f. elata was the 3rd layer > the 2nd layer > the 1st layer. In six treatments, combination of southern direction-the 3rd layer was the best technology of cultivation of Gastrodia elata f. elata, which had the best growth condition, the highest yield significantly higher than other treatments, and the best economic benefits. CONCLUSION: Southern direction associated with the 3rd layer is the best combination to planting Gastrodia elata f. elata in bionic wild cultivation. The planting ways not only improve the yield and quality, but also save land.

[Research on life history and phenological period of wild-stimulated cultivated Gastrodia elata f. elata in Guizhou].

In order to get to know the imitation of wild Gastrodia elata in life history and phenology period, by G. elata f. elata forest wild simulated cultivation in Dafang county, Guizhou province, observing and recording its morphological characteristics of each growth and development stage. This experiment summarized the law of its life history over 24 months, amplified the characteristics of each 5 phenology periods over the sexual and asexual reproduction of wild simulated cultivated G. elata f. elata in Guizhou. Which the results could clear the process of wild simulated cultivated G. elata f. elata in Guizhou, and provide a theoretical support for the standard technical of the simulated wild G. elata.

[The degradation on cultivated Gastrodia populations and prevention measures].

OBJECTIVE: To propose views and concepts on the degradation of cultivated Gastrodia populations. METHODS: The differences between natural and cultivated Gastrodia populations were compared and discussed in terms of their biological features, qualities and yields. RESULTS: There were changes and degradation in biological features and some important properties of yield in cultivated Gastrodia. CONCLUSION: The direct reason of cultivated Gastrodia degradation is the ecological disturbace of forests and the decreasing amount of natural Gastrodia populations; The indirect reason is manual pollination and other changes of the natural growing environment.

[The determination of three effective constituents in wild and cultivated Gastrodia elata from Bomi].

OBJECTIVE: To determine the contents of gastrodin, amino acids and total flavonoids in wild and cultivated Gastrodia elata, in order to choose the best method for its cultivation. The Gastrodia elata was picked at Guxiang town Bomi county Linzhi Region in Tibet. METHODS: HPLC was used to determine the content of gastrodin. The autoanalyzer was used to determine the content of amino acid. The ultraviolet spectrophotometer was adopted to measure the content of total flavonoids. RESULTS: The Gastrodin in wild Gastrodia elata was the highest. The contents of amino acids and total flavonoids in organic cultivated Gastrodia elata were higher than those in common cultivated and wild Gastrodia elata. CONCLUSION: The organic cultivated Gastrodia elata has better quality.

Relationships among genetic makeup, active ingredient content, and place of origin of the medicinal plant Gastrodia tuber.

Gastrodia tuber and its component gastrodin have many pharmacological effects. The chemical fingerprints and gastrodin contents of eight Gastrodia populations were determined, and the genomic DNA polymorphism of the populations was investigated. Genetic distance coefficients among the populations were calculated using the DNA polymorphism data. A dendrogram of the genetic similarities between the populations was constructed using the genetic distance coefficients. The results indicated that the genomic DNA of Gastrodia tubers was highly polymorphic; the eight populations clustered into three major groups, and the gastrodin content varied greatly among these groups. There were obvious correlations among genetic makeup, gastrodin content, and place of origin. The ecological environments in Guizhou and Shanxi may be conducive to evolution and to gastrodin biosynthesis, and more suitable for cultivation of Gastrodia tubers. These findings may provide a scientific basis for overall genetic resource management and for the selection of locations for cultivating Gastrodia tubers.

[Effects of drought on output of Gastrodia elata].

The experiments of water supply showed out drought can decrease water potencial and the output of Gastrodia elata. Those negative effects might be conpensated by hybridi combination, and its output should be increased.

[RAPD analysis on the germplasm resources of Gastrodia elata in Guizhou].

Genetic polymorphisms of genomic DNA of 15 samples from wild and culturaled Gastrodia elata Blume in Guizhou were analyzed by RAPD method. 12 effective primers are screened from 40 primers amplified a total number of 93 loci, among which 66 are polymorphic and the percentage of polymorphic loci (PPB) is 70.97%. UPGMA dendrogram analyzed by NTSYSpc, ver. 2.2 shows that Gastrodia elata have apparent genetic variance. Geological distribution and growing environment were significant factors for the polymorphism.

[Studies on Armillariella mellea cultivation with different kinds of trees].

The branches of several common trees in north China were selected to cultivate Armillariella mellea. The rhizomorph and growth speed of Armillariella mellea were measured,and the extracellular laccase activity was determined. The results showed that all tested trees except Sophora japonica and Amorpha fruiticosa were suitable to the growth of Armillariella mellea.

[Isozyme analysis of Gastrodia elata f. elata and G. elata f. glaucca and their hybrid].

Peroxidase(POX) isozyme analysis of Gastrodia elata f. glaucca, G. elata f. elata and its hybrid were carried out. The results indicated that shoot apex meristem was optimal material for POX analysis and the determination of the hybrid. The zymogram hands were stable and clear. POX isozyme zymogram of the hybrid F1(H x W) represented the visible supplement relation between Gastrodia elata f. elata and Gastrodia elata f. glaucca.

[The changes of cell structure in the courses of Armillaria mellea penetrating the nutritional stems of Gastrodia elata].

OBJECTIVE: To study the cell structure changes of Gastrodia elata after Armillaria mellea infection as well as the nutritional resource of this important medicinal plant. METHOD: Observation of the serial sections on the nutritional stems of G. elata, and measure new corms of G. elata when cut off the old corms connection with rhizomorph of A. mellea. RESULT: After G. elata nutritional stems infected by A. mellea, the rhizomorph of A. mellea separate into several hyphal layers and penetrate hyphal stream which infect into the cells of cortex layer in the direction of outside and infect directly into layer cells in the direction of inside, these hypha can be used the nutrition of G. elata the new corms will be stop growth if cut off the rhizomorph of A. mellea connected with G. elata. CONCLUSION: After the rhizomorph of A. mellea infected G. elata nutritional stems, hyphal coils and the hyphal stream will breakthrough passage cells as well as large cells surround the whole nutritional stems, so the passage cells is the key nutritional resource in the whole growth period of G. elata.

[Habitats and present situations of 13 rare precious medicinal materials in Funiu mountains].

The living resources are very abound in the Funiu mountains area because of its special geographical position and climate, and there are also many rare precious living beings and Chinese medicinal herbs, for example, Taxus mairei, Ginkgo biloba, Gastrodia elata, Dendrobium huashanense, Physochlaina infunbularis, Cypripedium japonicum, Dysosma versipellis, Moschus moschiferus, Megalobatrachus davidanus, Succinum, etc, and conscious protection and rational development is an urgent task.