Protoilludane-type sesquiterpenoids from Armillaria sp. by co-culture with the endophytic fungus Epicoccumsp. associated with Gastrodia elata.

An investigation of a co-culture of the Armillaria sp. and endophytic fungus Epicoccum sp. YUD17002 associated with Gastrodia elata led to the isolation of eight new compounds, including five protoilludane-type sesquiterpenes (1-5) and three aryl esters (6-8), together with six known analogues (9-14). The assignments of their structures were conducted via extensive analyses of the spectroscopic data and comparison of experimental and calculatedelectronic circular dichroism(ECD)data. Notably, these new compounds were not present in the pure culture controls and were only detected in the co-cultures. Compound 4 is the first example of an ent-protoilludane sesquiterpenoid scaffold bearing a five-membered lactone. Compound 6 exhibited moderate in vitro cytotoxic activities against five human cancer cell lines (HL-60, A549, MCF-7, SMMC-7721, and SW480) with IC50 values ranging from 15.80 to 23.03 µM. Moreover, 6 showed weak acetylcholinesterase inhibitory activity (IC50 value of 23.85 µM).

[Identification of four Armillaria strains and their effects on quality and yield of Gastrodia elata f. glauca].

In order to improve the quality and yield of Gastrodia elata f. glauca,determine the suitable Armillaria strains for the accompanying experiment in Xiaocaoba,Yiliang,four Armillaria strains were selected. They were used for G. elata cultivation,and the gene sequence,r DNA-ITS,ß-tubulin and EF1-α of four Armillaria strains,were compared and analyzed. The yield was mesured in November which was based on previous laboratory research. The tubers were washed and steamed,then dried and powdered. The content of gastrodin and p-hydroxybenzyl alcohol was determined by UPLC,the polysaccharide was determined by phenol-concentrated sulfuric acid method. The results showed that the strains M1,M2,M3 and M4 were Armillaria gallica group but there were differences in the yield and active ingredient content when they were cultivated with the same G. elata. The yield of G. elata( Jian Ma) was the lowest when cultivated with Armillaria strain M3,but it was not the same when used M1,0. 981 kg·m-2,the highest yield in the four stains.The content of gastrodin was 0. 581%,the total content of gastrodin and p-hydroxybenzyl alcohol was 0. 595%,when accompanied with M1 strains. It was higher than other strains. The content of G. elata polysaccharide was 2. 132%,which was similar to the content of M3 strain,higher than that of M2 and M4 strain. Selecting phylogenesis of Armillaria strians,the content of active ingredient,and the yield as indicators,it was concluded concerned that the M1 strain was the best of four strains. The results will provide a theoretical basis and guidance for higher yield and quality in cultivation of G. elata in Yiliang.

Hallmarks of Comparative Transcriptome between Rhizomorphs and Hyphae of Armillaria sp. 541 Participating in Fungal Symbiosis with Emphasis on LysM Domains.

Armillaria sp. 541, a genus of root-infecting fungi, forms a symbiosis with traditional Chinese medicine Gastrodia elata (Orchid) and Polyporus umbellatus via extensive networks of durable rhizomorphs. It is not clear the hallmarks of comparative transcriptome between the rhizomorphs and hyphae of Armillaria sp. 541. In the present study, transcriptomic analysis of Armillaria sp. 541 identified 475 differentially expressed genes (DEGs) between Armillaria rhizomorphs (AR) and hyphae (AH). Of them, 285 genes were upregulated and 190 were downregulated. Bioinformatics analyses and tests demonstrated DEGs involved in oxidoreductase activity and peptidoglycan binding were significantly enriched in this process when rhizomorph formed from hyphae. We accordingly obtained 14 gene-encoding proteins containing the LysM domain, and further consensus pattern and phylogenetic analysis indicated that their amino acid sequences were conserved and their biological functions may be peptidoglycan binding for recognition between the fungus and host. Among these genes, one, named Armillaria LysM domain recognition gene (aLDRG), was expressed significantly when rhizomorphs were differentiated from hyphae. It was located in the cortical cells of the rhizomorph by in situ hybridization. Furthermore, biolayer interferometry (BLI) assay demonstrated that aLDRG can bind specifically to chitin oligosaccharide of the fungal cell wall, including N,N',N″-Triacetylchitotriose (CO3) and N,N',N″,N'″,N″″-Pentaacetylchitopentaose (CO5). Therefore, we deduced that Armillaria sp. 541 expressed higher levels of LysM protein aLDRG for better binding of oligosaccharide after rhizomorphs were generated. This study provides functional genes for further studies on the interaction between Armillaria sp. 541 and its host.

Identification of four Armillaria strains and their effects on quality and yield of Gastrodia elata f. glauca / 中国中药杂志

In order to improve the quality and yield of Gastrodia elata f. glauca,determine the suitable Armillaria strains for the accompanying experiment in Xiaocaoba,Yiliang,four Armillaria strains were selected. They were used for G. elata cultivation,and the gene sequence,r DNA-ITS,β-tubulin and EF1-α of four Armillaria strains,were compared and analyzed. The yield was mesured in November which was based on previous laboratory research. The tubers were washed and steamed,then dried and powdered. The content of gastrodin and p-hydroxybenzyl alcohol was determined by UPLC,the polysaccharide was determined by phenol-concentrated sulfuric acid method. The results showed that the strains M1,M2,M3 and M4 were Armillaria gallica group but there were differences in the yield and active ingredient content when they were cultivated with the same G. elata. The yield of G. elata( Jian Ma) was the lowest when cultivated with Armillaria strain M3,but it was not the same when used M1,0. 981 kg·m-2,the highest yield in the four stains.The content of gastrodin was 0. 581%,the total content of gastrodin and p-hydroxybenzyl alcohol was 0. 595%,when accompanied with M1 strains. It was higher than other strains. The content of G. elata polysaccharide was 2. 132%,which was similar to the content of M3 strain,higher than that of M2 and M4 strain. Selecting phylogenesis of Armillaria strians,the content of active ingredient,and the yield as indicators,it was concluded concerned that the M1 strain was the best of four strains. The results will provide a theoretical basis and guidance for higher yield and quality in cultivation of G. elata in Yiliang.