Amanita Species Research Articles

An acid-tolerant lectin coupled with high Hg2+ potentiated hemagglutination enhancing property purified from Amanita hemibapha var. ochracea

A 37.4kDa acid tolerant lectin was isolated and purified from dried fruiting bodies of Amanita hemibapha var. ochracea designated as AHL. The lectin was not adsorbed on DEAE-cellulose, but rather adsorbed on S-Sepharose and subjected to gel filtration by fast protein liquid chromatography on Superdex 75. The purified lectin was immune from inhibition activities of metal ions. More over, AHL exhibited high agglutination activity on rabbit erythrocytes with accelerating Hg2+ ions concentration. Partial peptide sequence analysis (VSNNLLTGPKVVR) of this lectin showed relative similarity to phosphoenolpyruvate carboxykinase [ATP]-like protein as predicted from Fragaria vesca subsp. Vesca. Interestingly, AHL displayed a strong affinity toward α-Lactose, making our study the first report associating Amanita species’ lectin specificity for α-Lactose to the best of our knowledge.

Taxonomic Study of Amanita Subgenus Lepidella and Three Unrecorded Amanita Species in Korea

Amanita Pers. is a well-known monophyletic mushroom genus with a broad distribution. However, the diversity of Korean Amanita species has been underestimated, and most taxonomic studies conducted in Korea have only investigated their morphological characteristics. This approach is frequently insufficient for correct identification in fungal classification; therefore, we constructed a phylogeny of Amanita subgen. Lepidella in order to understand the phylogenetic placements of 16 Amanita specimens collected in Korea in 2012. The phylogeny constructed using the sequence data of the internal transcribed spacers and the partial large subunit of ribosomal RNA identified nine Amanita species (A. citrina, A. excelsa var. spissa, A. flavipes, A. fritillaria, A. oberwinklerana, A. pallidorosea, A. rubescens, A. subjunquillea, and A. volvata); of these, A. fritillaria, A. oberwinklerana, and A. pallidorosea are new to Korea.

Development, optimization and application of an analytical methodology by ultra performance liquid chromatography–tandem mass spectrometry for determination of amanitins in urine and liver samples

Amanitins, highly toxic cyclopeptides isolated from various Amanita species, are the most potent poisons accounting for the hazardous effects on intestinal epithelium cells and hepatocytes, and probably the sole cause of fatal human poisoning.The present study was focused on the development, optimization and application of an analytical methodology by ultra performance liquid chromatography-tandem mass spectrometry (UPLC–MS/MS), following urine and liver sample preparation by protein precipitation with organic solvents, and solid phase extraction (SPE) procedure, for the determination of the amatoxins, α- and β-amanitin. Linearity, detection and quantification limits, selectivity, sensitivity, intra and inter-assay precision and recovery were studied, in order to guarantee reliability in the analytical results. The developed method proved to be specific and selective, with LOD (Limit of Detection) values for α- and β-amanitin of 0.22 and 0.20ngmL−1 in urine and 10.9 and 9.7ngg−1 in liver, respectively. LOQ (Limit of Quantification) values ranged from 0.46 to 0.57ngmL−1in urine and 12.3–14.7ngg−1 in tissue, for both amanitins. Linearity, in the range of 10.0–200.0ngmL−1or ngg−1, shows that coefficients of correlation were greater than 0.997 for α-amanitin and 0.993 for β-amanitin. Precision was checked at three levels during three consecutive days with intra-day and inter-day coefficients of variation not greater than 15.2%. The extraction recovery presents good results for the concentrations analyzed, with values ranging from 90.2 to 112.9% for both matrices.Thus, the proposed analytical method is innovative, presents a high potential in the identification, detection and determination of α- and β-amanitins in urine and tissue samples, as well as in other biological samples, such as kidney and mushrooms.

Four Newly Recorded Amanita Species in Korea: Amanita sect. Amanita and sect. Vaginatae

We collected nearly 70 specimens of Amanita species during a diversity study of Korean mushrooms conducted in 2012. In this study, we primarily investigated 23 Amanita specimens belonging to sections Amanita and Vaginatae. Based on sequence data of the internal transcribed spacers and partial large subunit of ribosomal RNA and morphological characteristics, we identified the following 15 phylogenetic species: A. alboflavescens, A. ceciliae, A. farinosa, A. fulva, A. griseofolia, A. ibotengutake, A. melleiceps, A. orientifulva, A. pantherina, A. rubrovolvata, A. sinensis, A. subglobosa, A. vaginata, A. cf. vaginata f. alba, and an undescribed Amanita species. In this study, four of the identified Amanita species (A. griseofolia, A. ibotengutake, A. orientifulva, and A. sinensis) were reported for the first time in Korea.

Investigation and analysis of 102 mushroom poisoning cases in Southern China from 1994 to 2012

Mushroom poisoning is the main cause of mortality in food poisoning incidents in China. Although some responsible mushroom species have been identified, some were identified inaccuratly. This study investigated and analyzed 102 mushroom poisoning cases in southern China from 1994 to 2012, which involved 852 patients and 183 deaths, with an overall mortality of 21.48 %. The results showed that 85.3 % of poisoning cases occurred from June to September, and involved 16 species of poisonous mushroom: Amanita species (A. fuliginea, A. exitialis, A. subjunquillea var. alba, A. cf. pseudoporphyria, A. kotohiraensis, A. neoovoidea, A. gymnopus), Galerina sulciceps, Psilocybe samuiensis, Russula subnigricans, R. senecis, R. japonica, Chlorophyllum molybdites, Paxillus involutus, Leucocoprinus cepaestipes and Pulveroboletus ravenelii. Six species (A. subjunquillea var. alba, A. cf. pseudoporphyria, A. gymnopus, R. japonica, Psilocybe samuiensis and Paxillus involutus) are reported for the first time in poisoning reports from China. Psilocybe samuiensis is a newly recorded species in China. The genus Amanita was responsible for 70.49 % of fatalities; the main lethal species were A. fuliginea and A. exitialis. Russula subnigricans caused 24.59 % of fatalities, and five species showed mortality >20 % (A. fuliginea, A. exitialis, A. subjunquillea var. alba, R. subnigricans and Paxillus involutus). Mushroom poisoning symptoms were classified from among the reported clinical symptoms. Seven types of mushroom poisoning symptoms were identified for clinical diagnosis and treatment in China, including gastroenteritis, acute liver failure, acute renal failure, psychoneurological disorder, hemolysis, rhabdomyolysis and photosensitive dermatitis.

In vitro mycorrhization and acclimatization of Amanita caesareoides and its relatives on Pinus densiflora

Amanita caesareoides is a sister species of Amanita caesarea, also known as Caesar's mushroom and one of the most desirable edible mycorrhizal mushrooms. However, cultivation of Caesar's mushrooms has not yet been successful due to the difficulties involved in establishing pure cultures. In this study, we established pure cultures of four Asian Caesar's mushroom species, i.e., A. caesareoides, Amanita javanica, Amanita esculenta, and Amanita similis, which were identified by sequence analysis of their rDNA internal transcribed spacer (ITS) region. Five selected isolates in A. caesareoides, A. javanica, and A. esculenta were tested for ectomycorrhizal syntheses with axenic Pinus densiflora seedlings in vitro. Ectomycorrhizal tips of each fungal isolate tested were observed on pine lateral roots within 5 months of inoculation. Seventeen pine seedlings that formed ectomycorrhizas in vitro with these three Amanita species were acclimatized under non-sterile conditions. Seven months following acclimatization, ectomycorrhizal colonization by A. caesareoides was observed on newly grown root tips, which was confirmed by polymerase chain reaction restriction fragment length polymorphism analysis of the fungal rDNA ITS region. Two other Amanita species also survived during ectomycorrhizal acclimatization. These results suggest that the cultivation of A. caesareoides and its relatives can be attempted through mycorrhizal synthesis using P. densiflora as a host. This is the first report of in vitro mycorrhization of Asian Caesar's mushrooms and their acclimatization under non-sterile conditions.

The Irreversible Loss of a Decomposition Pathway Marks the Single Origin of an Ectomycorrhizal Symbiosis

Microbial symbioses have evolved repeatedly across the tree of life, but the genetic changes underlying transitions to symbiosis are largely unknown, especially for eukaryotic microbial symbionts. We used the genus Amanita, an iconic group of mushroom-forming fungi engaged in ectomycorrhizal symbioses with plants, to identify both the origins and potential genetic changes maintaining the stability of this mutualism. A multi-gene phylogeny reveals one origin of the symbiosis within Amanita, with a single transition from saprotrophic decomposition of dead organic matter to biotrophic dependence on host plants for carbon. Associated with this transition are the losses of two cellulase genes, each of which plays a critical role in extracellular decomposition of organic matter. However a third gene, which acts at later stages in cellulose decomposition, is retained by many, but not all, ectomycorrhizal species. Experiments confirm that symbiotic Amanita species have lost the ability to grow on complex organic matter and have therefore lost the capacity to live in forest soils without carbon supplied by a host plant. Irreversible losses of decomposition pathways are likely to play key roles in the evolutionary stability of these ubiquitous mutualisms.

Amatoxin Poisoning: Case Reports and Review of Current Therapies

BackgroundDiagnosis and management of Amanita mushroom poisoning is a challenging problem for physicians across the United States. With 5902 mushroom exposures and two resultant deaths directly linked to Amanita ingestion in 2009, it is difficult for physicians to determine which patients are at risk for lethal toxicity. Identification of amatoxin poisoning can prove to be difficult due to delay in onset of symptoms and difficulty with identification of mushrooms. Consequently, it is difficult for the Emergency Physician to determine proper disposition. Further, treatment options are controversial. ObjectivesTo review current data to help health care providers effectively identify and treat potentially deadly Amanita mushroom ingestions. Case ReportsWe present two cases of Amanita mushroom ingestion in the northeastern United States treated with N-acetylcysteine, high-dose penicillin, cimetidine, and silibinin, a semi-purified fraction of milk thistle-derived silymarin, as part of their treatment regimen. The mushroom species was identified by a consultant as Amanita Ocreata. ConclusionsWe present the successful treatment of 2 patients who ingested what we believe to be an Amanita species never before identified in the northeastern United States.

Amanita thiersii is a saprotrophic fungus expanding its range in the United States

Although most species in the genus Amanita form ectomycorrhizal associations, a few are reported to be saprotrophs living in grassland habitats. Little is known about the ecology and distribution of these free-living Amanita species. We describe the ecology of Amanita thiersii, a species commonly collected in lawns throughout the Mississippi River Basin. Stable isotopes of carbon, transcriptomic sequences and patterns of growth on complex carbon sources provide evidence for A. thiersii as a saprotrophic species. Sporocarps of A. thiersii are less depleted in 13C compared to published data for ectomycorrhizal fungi, supporting a saprotrophic mode of carbon acquisition in the field. Orthologs of cellulase genes known to play key roles in the decomposition of cellulose in other basidiomycetes were identified in a transcriptome of A. thiersii, establishing that this species has the genetic potential to degrade cellulose. Amanita thiersii also can use artificial cellulose or sterile grass litter as a sole carbon source. DNA sequences of three nuclear gene regions and banding patterns from four inter-simple sequence repeat markers were identical across 31 populations from throughout the known range of the species, which suggests the genetic diversity of A. thiersii populations is low. Maps of A. thiersii collections made from the 1950s until present suggest this species is experiencing a range expansion. It was reported first in 1952 in Texas and now occurs in nine states north to Illinois. These data provide an ecological context for interpreting the genome of A. thiersii, currently being sequenced at the United States Department of Energy’s Joint Genome Institute.

Ribosomal biosynthesis of α-amanitin in Galerina marginata

Amatoxins, including α-amanitin, are bicyclic octapeptides found in mushrooms (Agaricomycetes, Agaricales) of certain species in the genera Amanita, Galerina, Lepiota, and Conocybe. Amatoxins and the chemically similar phallotoxins are synthesized on ribosomes in Amanita bisporigera, Amanita phalloides, and Amanita ocreata. In order to determine if amatoxins are synthesized by a similar mechanism in another, distantly related mushroom, we obtained genome survey sequence data from a monokaryotic isolate of Galerinamarginata, which produces α-amanitin. The genome of G. marginata contains two copies of the α-amanitin gene (GmAMA1-1 and GmAMA1-2). The α-amanitin proprotein sequences of G. marginata (35 amino acids) are highly divergent from AMA1 of A. bisporigera except for the toxin region itself (IWGIGCNP in single-letter amino acid code) and the amino acids immediately upstream (N[A/S]TRLP). G. marginata does not contain any related toxin-encoding sequences besides GmAMA1-1 and GmAMA1-2. DNA from two other α-amanitin-producing isolates of Galerina (G. badipes and G. venenata) hybridized to GmAMA1, whereas DNA from the toxin non-producing species Galerinahybrida did not. Expression of the GmAMA1 genes was induced by growth on low carbon. RNASeq evidence indicates that both copies of GmAMA1 are expressed approximately equally. A prolyl oligopeptidase (POP) is strongly implicated in processing of the cyclic peptide toxins of A. bisporigera and Conocybe apala. G. marginata has two predicted POP genes; one, like AbPOPB of A. bisporigera, is present only in the toxin-producing isolates of Galerina and the other, like AbPOPA of A. bisporigera, is present in all species. Our results indicate that G.marginata biosynthesizes amatoxins on ribosomes by a pathway similar to Amanita species, involving a genetically encoded proprotein of 35 amino acids that is post-translationally processed by a POP. However, due to the high degree of divergence, the evolutionary relationship between AMA1 in the genera Amanita and Galerina is unclear.

Amanita poisonings resulting in acute, reversible renal failure: new cases, new toxic Amanita mushrooms

Renal failure as a consequence of eating mushrooms has been reported repeatedly after ingestion of webcaps of the Cortinarius orellanus group. But mushrooms of the genus Amanita can also cause renal failure: Amanita smithiana (North America) and Amanita proxima (Mediterranean area). Here, we discuss poisonings caused by other white amanitas. A German and--independently--two Portuguese patients reported the ingestion of completely white mushrooms with ring. Similar to intoxications with A. smithiana or A. proxima, the clinical picture was characterized by nausea and vomiting 10-12 h after ingestion, severe acute renal failure and mild hepatitis. Renal biopsy showed acute interstitial nephritis and tubular necrosis. Two patients were given temporary haemodialysis. All have fully recovered their renal function. Poisonings caused by mushrooms containing the toxin of A. smithiana were suspected. We tested 20 Amanita species for the presence of this toxin. Thin layer chromatography was applied to detect A. smithiana nephrotoxin in herbarium specimens using authentic material of A. smithiana as reference. A. smithiana toxin could be detected in Amanita boudieri, Amanita gracilior and in Amanita echinocephala. A. boudieri was collected by the Portuguese patients. A. echinocephala is the only nephrotoxic Amanita growing North of the Alps and is suspected to be the cause of renal failure in the German patient. No A. smithiana toxin was detectable in the nephrotoxic A. proxima. A. boudieri, A. gracilior and A. echinocephala are nephrotoxic. These intoxications are clinically similar to that of A. smithiana, with acute reversible renal failure and mild hepatitis but are different in their clinical picture from Orellanus syndrome characterized by a delayed onset of severe and often irreversible renal failure.

Amatoxin and phallotoxin composition in species of the genus Amanita in Colombia: A taxonomic perspective

Some species in the genus Amanita have a great variety of toxic secondary metabolites. They are characterized macroscopically by having a white spore print and free gills, and microscopically by the presence of a divergent hymenophoral trama. Some species of Amanita present in Colombia were chemically characterized by analyzing their toxin composition using HPLC. Samples were collected in oak (Quercus humboldtii) and pine (Pinus radiata) forests. Twelve species were recovered, Amanita fuligineodisca, Amanita xylinivolva, Amanita flavoconia, Amanita rubescens, Amanita bisporigera, Amanita muscaria, Amanita humboldtii, Amanita sororcula, Amanita brunneolocularis, Amanita colombiana, Amanita citrina, Amanita porphyria as well as two unreported species. Results showed that most of the analyzed species have α -amanitin in concentrations ranging from 50 ppm to 6000 ppm. Concentrations of α-amanitin in the pileus were significantly greater than in the stipe. Phalloidin and phallacidin were only present in A. bisporigera. Chromatographic profiles are proposed as an additional taxonomic tool since specific peaks with similar retention times were conserved at the species level.

Development of a DNA-Based Macroarray for the Detection and Identification of Amanita Species

A DNA-based macroarray was designed to quickly and accurately identify certain Amanita mushroom specimens at the species level. The macroarray included probes for Amanita phalloides and Amanita ocreata, toxic species responsible for most mushroom poisonings, and Amanita lanei and Amanita velosa, edible species sometimes confused with toxic species, based on sequences of the highly variable internal transcribed spacer (ITS) region of rDNA. A cryptic species related to A. ocreata and one related to A. lanei, identifiable by ITS sequences, were also included. Specific multiple oligonucleotide probes were spotted onto nylon membranes and the optimal hybridization temperatures were determined. The Amanita DNA array was highly specific, sensitive (0.5 ng DNA/μL and higher were detected), and reproducible. In two case studies, the method proved useful when only small amounts of mushroom tissue remained after a suspected poisoning. An identification could be completed in 12 h.

Two rare white forms of common Amanita species

Two rare white forms of common Amanita species

Amatoxin and phallotoxin concentration in Amanita bisporigera spores

Even though amatoxins and phallotoxins have been well characterized in basidiocarps of Amanita species, to our knowledge no report of these toxins in spores of Amanitas has been published. Reversed phase HPLC was used to determine nonzero concentrations of α-amanitin (0.30 mg/g), and phallacidin (0.02 mg/g) in spores taken from white Amanita sect. phalloideae species. We did not find significant amounts of phalloidin in Amanita spores. We also report concentrations of these toxins from pileus and stipe tissues that are similar to previously reported values, lending support to the hypothesis that toxin concentrations in spores are much less than in other basidiocarp tissues.

Lethal amanitas of East Asia characterized by morphological and molecular data

This study investigated the morphological characteristics and molecular phylogenetic relationships among lethal Amanita species occurring in East Asia. The results revealed that at least nine phylogenetic taxa exist in the region. Among them, five were identical to previously known morphological taxa including A. exitialis, A. fuliginea, A. subjunquillea, A. subjunquillea var. alba and A. virosa; three new taxa, A. fuligineoides, A. rimosa and A. pallidorosea were described and illustrated here. The remaining one was labeled “A. sp. 1” because no robust morphological evidence was found to distinguish it from A. subjunquillea var. alba. The occurrence of A. virosa in East Asia, a well-known lethally poisonous mushroom originally described from Europe, was confirmed by both ITS sequences and morphology. Another lethal species native to Europe, A. phalloides, was determined as closely related to A. subjunquillea from East Asia. Amanita oberwinklerana was treated in section Phalloideae from a morphological point of view, but appeared to be a member of section Lepidella by the analyses of sequences from both the ITS regions and the large subunit of nuclear ribosomal RNA gene. Distribution features of East Asian Phalloideae and the phylogenetic relationships of these species with their counterparts from Europe and North America were also discussed. A key to species of section Phalloideae in East Asia is furnished.

Amanita viscidolutea, a new species from Brazil with a key to Central and South American species of Amanita section Amanita

We described and illustrated Amanita viscidolutea sp. nov. from specimens collected in the state of Rio Grande do Norte, northeastern Brazil. The main characteristics of the new species are its yellow pileus with white margin, the viscidity of the pileal surface, an exannulate stipe and inamyloid basidiospores. We also present an artificial dichotomous key to Central and South American species of Amanita (subgenus Amanita) section Amanita.

The ectomycorrhizal fungus Amanita phalloides was introduced and is expanding its range on the west coast of North America

The deadly poisonous Amanita phalloides is common along the west coast of North America. Death cap mushrooms are especially abundant in habitats around the San Francisco Bay, California, but the species grows as far south as Los Angeles County and north to Vancouver Island, Canada. At different times, various authors have considered the species as either native or introduced, and the question of whether A. phalloides is an invasive species remains unanswered. We developed four novel loci and used these in combination with the EF1α and IGS loci to explore the phylogeography of the species. The data provide strong evidence for a European origin of North American populations. Genetic diversity is generally greater in European vs. North American populations, suggestive of a genetic bottleneck; polymorphic sites of at least two loci are only polymorphic within Europe although the number of individuals sampled from Europe was half the number sampled from North America. Endemic alleles are not a feature of North American populations, although alleles unique to different parts of Europe were common and were discovered in Scandinavian, mainland French, and Corsican individuals. Many of these endemic European haplotypes were found together at single sites in California. Early collections of A. phalloides dated prior to 1963 and annotated using sequences of the ITS locus proved to be different species of Amanita. The first Californian collections that we confirmed as A. phalloides were made from the Del Monte Hotel (now the Naval Postgraduate School) in Monterey, and on the campus of the University of California, Berkeley, in 1938 and in 1945. These historical data are used in combination with data on A. phalloides' current distribution to estimate a rate of spread for A. phalloides in California. Many species of ectomycorrhizal (EM) fungi have been introduced across and among continents, but with this evidence, the death cap becomes the only known invasive EM fungus in North America.

Hallucinogenic Mushrooms in Mexico: An Overview

Hallucinogenic Mushrooms in Mexico: An Overview.Psilocybe, with 53 known hallucinogenic species in Mexico, is the most important and diverse group of sacred mushrooms used by Mexican indigenous cultures. Psilocybe caerulescens, known by the present-day Nahuatl Indians as teotlaquilnanacatl, is hypothesized to be the ceremonially-used teonanacatl mushroom cited by Sahagun in the 16th century, the true identity of which has remained obscure for centuries. Correcting a widely disseminated error derived from early published information on Mexican hallucinogenic mushrooms, emphasis is placed on the fact that Panaeolus species have never been used traditionally in Mexico. Reports of the use of species of Amanita, Clavaria, Conocybe, Cordyceps, Dictyophora, Elaphomyces, Gomphus, Lycoperdon, Psathyrella, and Stropharia as sacred or narcotic mushrooms are discussed. A brief history of the discovery of hallucinogenic mushrooms in Mexico is presented, as well as notes on their taxonomy, distribution, and traditional use in Mexico.

Identification of toxic oligopeptides in Amanita fungi employing capillary electrophoresis‐electrospray ionization‐mass spectrometry with positive and negative ion detection

The identification of toxic oligopeptides employing CE-ESI-MS is presented. The analytes studied ama- and phallotoxins are of significant forensic interest because over 90% of the lethal cases of fungus poisoning in man are caused by species of Amanita which contain these toxins. A CE method was developed to separate the toxins alpha-, beta- and gamma-amanitin, phalloidin and phallacidin. Their fragmentation patterns in MS(n) experiments were investigated in the positive and in the negative ion mode, also the influence of the sheath liquid mixture of the used interface on the S/N. Method validation included the determination of the LOD and the repeatability of the migration time and peak area for both detection modes. With the optimized method LODs of 13-79 ng/mL (17-87 nmol/L) were reached. The CE-MS procedure was successfully applied to the identification of ama- and phallotoxins in extracts of air-dried mushroom samples.