Two highly conjugated ergosterols from the fungus Psathyrella rogueiana and their anti-inflammatory activity.

Two previously undescribed ergosterols containing a highly conjugated ring system, psathrosterols A and B (1 and 2), have been isolated from the fungus Psathyrella rogueiana. Their structures with absolute configurations were established by extensive spectroscopic methods, as well as single crystal X-ray diffraction. Compounds 1 and 2 showed inhibitory activity against NO production with IC50 values of 22.3 and 16.4 µM, respectively.

Morphology and Molecular Phylogeny Reveal Five New Species of Laccaria (Hydnangiaceae, Agaricales) from Southern China.

The genus Laccaria is a type of cosmopolitan and ecologically important fungal group. Members can form ectomycorrhizal associations with numerous trees, and some species are common edible fungi in local markets. Although some new species from China are recently published, the species diversity of Laccaria is still unclear in China. In this study, some samples of Laccaria were collected from southern China, and morphological characteristics and phylogenetic analyses based on the multilocus dataset of ITS-LSU-tef1-rpb2 confirmed five new species. Laccaria miniata, L. nanlingensis and L. neovinaceoavellanea were collected from subtropical broad-leaved forests, and L. rufobrunnea and L. umbilicata were collected from subtropical mixed forests of southwest China. Full descriptions, illustrations, comparisons with similar species and phylogenetic analysis are provided.

New insights into the genus Gyroporus (Gyroporaceae, Boletales), with establishment of four new sections and description of five new species from China.

Species of Gyroporus from southern China were studied in this study. Based on morphology and molecular phylogenetic analyses of DNA sequences from the nuclear ribosomal internal transcribed spacer (ITS), the nuclear ribosomal large subunit (nrLSU), and the mitochondrial adenosine triphosphate ATP synthase subunit 6 (atp6), Gyroporus was divided into four main branches in the phylogenetic tree, and four sections were firstly proposed i.e. Gyroporus sect. Castaneus, G. sect. Cyanescens, G. sect. Longicystidiatus and G. sect. Pallidus. Five new species, i.e. G. alboluteus, G. atrocyanescens, G. pseudolongicystidiatus, G. pallidus and G. subcaerulescens, were revealed from China, and their phylogenetic positions were also analysed. Among them, G. alboluteus and G. pallidus were nested into the sect. Pallidus, although morphologically similar to G. castaneus; G. atrocyanescens and G. subcaerulescens, with obvious cyanescent oxidation reactions, were nested into the sect. Cyanescens; and G. pseudolongicystidiatus characterised by its long cystidia and was nested into the sect. Longicystidiatus. The new species were formally described and illustrated in the present study, and a key to the sections and species of Gyroporus in China was provided.

Diversity of Cantharellus (Cantharellales, Basidiomycota) in China with Description of Some New Species and New Records.

Cantharellus is a well-known genus of edible mushrooms, belonging to the family Hydnaceae in the class Agaricomycetes. In this study, a phylogenetic overview of Cantharellus subg. Cinnabarinus and C. subg. Parvocantharellus in China is carried out with the description of four new species. Species description are based on morphological characters of basidiomata and phylogenetic analyses of multi-locus dataset of 28S + tef1 + rpb2. Among the new species, two species, C. chrysanthus and C. sinocinnabarinus, belong to C. subg. Cinnabarinus and two new species, C. convexus and C. neopersicinus, belong to C. subg. Parvocantharellus. Species delimitation characters of the new taxa are compared with closely related species. In addition, three new records of Cantharellus are reported for China: C. albovenosus and C. citrinus of subg. Cinnabarinus and C. koreanus of subg. Parvocantharellus. A key to the species of subg. Cinnabarinus in China was provided.

Multigene Phylogeny and Morphology Reveal Unexpectedly High Number of New Species of Cantharellus Subgenus Parvocantharellus (Hydnaceae, Cantharellales) in China.

The genus Cantharellus, commonly known as chanterelles, has recently been divided into six subgenera; however, wider sampling approaches are needed to clarify the relationships within and between these groups. A phylogenetic overview of Cantharellus subgenus Parvocantharellus in China was inferred based on the large subunit nuclear ribosomal RNA gene (nrLSU), the DNA-directed RNA polymerase II subunit 2 (rpb2), and the transcription elongation factor 1-alpha (tef1). A total of nine species from China were assigned to the subgenus, including seven novel species, namely Cantharellusaurantinus, C. austrosinensis, C. galbanus, C. luteolus, C. luteovirens, C. minioalbus, and C. sinominior, and two known species, namely C. albus and C. zangii. The detailed descriptions and illustrations were provided based on the newly obtained data, with the comparisons to closely related species. C. zangii was restudied based on the paratype specimens and multiple new collections from the type locality. Futhermore, the Indian species C. sikkimensis was identified as a synonym of C. zangii based on the morphological and molecular analyses. A key to the Chinese species belonging to the subg. Parvocantharellus is also provided.

Two new species of Phallus (Phallaceae) with a white indusium from China.

Two new Phallus species, P.cremeo-ochraceus and P.rigidiindusiatus were discovered in southwestern and southern China, respectively. Phalluscremeo-ochraceus is morphologically characterized by its cream to ochraceous receptacle, white to very slightly pinkish indusium, white to pinkish pseudostipe and white to slightly purplish pink volva. Phallusrigidiindusiatus is characterized by a white to yellowish white receptacle, a strongly rigid indusium usually without serrated margin and smaller basidiospores than those of P.serratus. Phylogenetic positions of the two species are located in two independent lineages respectively. Detailed descriptions, color photographs, illustrations and a key to the related species are presented.

Two new Inosperma (Inocybaceae) species with unexpected muscarine contents from tropical China.

An accurate identification of poisonous mushrooms and the confirmation of the toxins involved are both of great importance in the treatment of mushroom poisoning incidents. In recent years, cases of mushroom poisoning by Inosperma spp. have been repeatedly reported from tropical Asia. It is urgent to know the real species diversity of Inosperma in this region. In the present study, we proposed two new Inosperma species from tropical Asia, namely I.muscarium and I.hainanense. They were described based on morphology and multilocus phylogeny. Detailed descriptions, color photographs and the discussion with other closely related species of the two new taxa were provided. In addition, a comprehensive muscarine determination of these two new species using ultrahigh performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) approach has been performed. Results showed that these two species were muscarine positive, with a content of 16.03 ± 1.23 g/kg in I.muscarium and a content of 11.87 ± 3.02 g/kg in I.hainanense, much higher than the known species I.virosum. Recovery of muscarine ranged from 93.45% to 97.25%, and the average recovery is 95.56%.

Transcriptome sequencing analysis of the MSDIN gene family encoding cyclic peptides in lethal Amanita fuligineoides.

Amanita fuligineoides, a lethal mushroom discovered in China, contains abundant cyclic peptide toxins that can cause fatal poisoning. However, the MSDIN gene family encoding for these cyclic peptides in A. fuligineoides has not been systematically studied. In this research, the transcriptome sequencing of A. fuligineoides was performed and its MSDIN family members were analyzed. A total of 4.41 Gb data containing 30833 unigenes was obtained; sequence alignments throughout several databases were done to obtain their functional annotations. Based on these annotations, MSDIN genes were found and verified by RT-PCR. A total of 29 different core peptides were obtained: 3 toxin genes, encoding ß-amanitin (ß-AMA), phalloidin (PHD), and phallacidin (PCD), and 26 genes encoding unknown cyclic peptides, 20 of which are reported for the first time and may encode for novel cyclic peptides. Analysis of the predicted precursor peptides indicated that octocyclic peptides were the main MSDIN peptides synthesized by A. fuligineoides, accounting for the 45%. A phylogenetic analysis suggested that studied precursor peptides could be clustered into 7 clades, which might represent different functionalities. Results suggested that A. fuligineoides might have a strong capacity to synthesize cyclopeptides, laying the foundation for their excavation and utilization.

Clitopilus, Clitocella, and Clitopilopsis in China.

Within the family Entolomataceae, many reports of Entoloma and Rhodocybe have been published, but few reports on Clitopilus and allied genera are available. In this study, Clitopilus, Clitocella, and Clitopilopsis from China were studied by morphological and phylogenetic methods. Portions of nuc 28S rDNA (28S), RNA polymerase II second largest subunit (rpb2), translation elongation factor 1-alpha (tef1), and ATPase subunit 6 (atp6), were employed to elucidate the relationships of Clitopilus and allies. Results indicate that Clitopilus should be divided into three sections, namely, sect. Clitopilus, sect. Scyphoides, and sect. Crispi, the last of which is newly described. In total, 17 species of Clitopilus, Clitocella, and Clitopilopsis are found in China; 14 species belong to Clitopilus, of which 5 are new: C. yunnanensis, C. brunniceps, C. rugosiceps, C. sinoapalus, and C. umbilicatus. Two species of Clitocella, including one new species, Clitocella orientalis, are described. One new species of Clitopilopsis, namely, Clitopilopsis albida, is proposed.

Phylogenetic overview of Aureoboletus (Boletaceae, Boletales), with descriptions of six new species from China.

In this study, species relationships of the genus Aureoboletus were studied, based on both morphological characteristics and a four-gene (nrLSU, tef1-a, rpb1 and rpb2) phylogenetic inference. Thirty-five species of the genus have been revealed worldwide, forming eight major clades in the phylogenetic tree, of which twenty-four species have been found in China, including six new species: A. glutinosus, A. griseorufescens, A. raphanaceus, A. sinobadius, A. solus, A. velutipes and a new combination A. miniatoaurantiacus (Bi & Loh) Ming Zhang, N.K. Zeng & T.H. Li proposed here. A key to 24 known Chinese species has been provided.

Molecular cloning and the expression pattern of AePOPB involved in the α-amanitin biosynthesis in Amanita exitialis fruiting bodies.

Amanita exitialis Zhu L. Yang & T. H. Li is the species responsible for the largest number of mushroom-associated human poisonings and fatalities in South China due to its lethal cyclic peptide toxins. Prolyl oligopeptidase B (POPB) is considered a key enzyme in the production of the highly toxic cyclic peptide α-amanitin. However, the POPB gene of A. exitialis has not been studied. In the present study we cloned and sequenced the full-length A. exitialis POPB (AePOPB) gene. The aim was to verify the gene structure and functions of AePOPB. The full-length sequence of AePOPB is 3144 bp, including 18 exons encoding 730 aa, and the advanced structure is very similar to that of the previously reported POPB in Galerina marginata (GmPOPB). The amino acid sequence of AePOPB is highly homologous with those from other amanitin-producing lethal mushrooms, implying that AePOPB may have a similar role in the biosynthesis of cyclic peptide toxins. Expression levels of AePOPB were detectable in all parts and developmental stages of the fruiting bodies, and AePOPB was expressed more strongly at early development stages (early and late elongation stages). At early and late elongation stages, the expression peaks occurred in the stipe, whereas at early and late mature stages, the expression peaks occurred in the pileus. The expression patterns of AePOPB in different stages and different parts of the fruiting bodies were highly consistent with those of Aeα-AMA, which is required for α-amanitin accumulation. These results indicate that AePOPB should be involved in the α-amanitin biosynthesis in A. exitialis.

Illumina-based de novo transcriptome sequencing and analysis of Amanita exitialis basidiocarps.

Amanita exitialis is a lethal mushroom that was first discovered in Guangdong Province, China. The high content of amanitin in its basidiocarps makes it lethal to humans. To comprehensively characterize the A. exitialis transcriptome and analyze the Amanita toxins as well as their related gene family, transcriptome sequencing of A. exitialis was performed using Illumina HiSeq 2000 technology. A total of 25,563,688 clean reads were collected and assembled into 62,137 cDNA contigs with an average length of 481 bp and N50 length of 788 bp. A total of 27,826 proteins and 39,661 unigenes were identified among the assembled contigs. All of the unigenes were classified into 166 functional categories for understanding the gene functions and regulation pathways. The genes contributing to toxic peptide biosynthesis were analyzed. From this set, eleven gene sequences encoding the toxins or related cyclic peptides were discovered in the transcriptome. Three of these sequences matched the peptide toxins α-amanitin, ß-amanitin, and phallacidin, while others matched amanexitide and seven matched unknown peptides. All of the genes encoding peptide toxins were confirmed by polymerase chain reaction (PCR) in A. exitialis, and the phylogenetic relationships among these proprotein sequences were discussed. The gene polymorphism and degeneracy of the toxin encoding sequences were found and analyzed. This study provides the first primary transcriptome of A. exitialis, which provided comprehensive gene expression information on the lethal amanitas at the transcriptional level, and could lay a strong foundation for functional genomics studies in those fungi.

Peptide toxin components of Amanita exitialis basidiocarps.

Eight peptide toxins were isolated and purified from basidiocarps of Amanita exitialis with high performance liquid chromatography and were subjected to ultraviolet, nuclear magnetic resonance and mass spectrometry. We identified seven peptide toxins, α-amanitin, ß-amanitin, amaninamide, phallacin, phallacidin, phallisacin and desoxoviroidin. The molecular weight (729.5 Da) of the eighth compound did not match that of any reported Amanita toxins and, although the UV absorption spectrum indicated it to be a phallotoxin, further studies are required to identify this component. This is the first report of amaninamide, phallacin, phallisacin and desoxoviroidin in this lethal mushroom species.

[Study on intraspecific genetic diversity in different plant populations of Pogostemon cabli].

OBJECTIVE: To study the genetic polymorphism and intraspecific genetic differentiation of different populations of Pogostemon cablin, and find out the effective method to distinguish DNA fingerprint of different populations of P. cablin. METHOD: Five plant populations of P. cablin were analyzed by RAPD markers. PopGen 32 software for clustering analysis and calculating. Fourteen of the 80 random primers were tested to possess the stronger detecting effect of polymorphous character. RESULT: A total of 84 bands was amplified by the 10 primers, among them 17 bands were monomorphic. 67 of them were polymorphic. The results indicated that the genetic variations existed within the different plant populations of the same species. CONCLUSION: It is feasible by RAPD technique with specifically primer to analyze the genetic diversity and identify 5 plant populations of P. cablin. RAPD technique has provided a new path for identification and classification of P. cablin genetic germplasm.