Identifying the “Mushroom of Immortality”: Assessing the Ganoderma Species Composition in Commercial Reishi Products

Species of Ganoderma, commonly called reishi (in Japan) or lingzhi (in China), have been used in traditional medicine for thousands of years, and their use has gained interest from pharmaceutical industries in recent years. Globally, the taxonomy of Ganoderma species is chaotic, and the taxon name Ganoderma lucidum has been used for most laccate (shiny) Ganoderma species. However, it is now known that G. lucidum sensu stricto has a limited native distribution in Europe and some parts of China. It is likely that differences in the quality and quantity of medicinally relevant chemicals occur among Ganoderma species. To determine what species are being sold in commercially available products, twenty manufactured products (e.g., pills, tablets, teas, etc.) and seventeen grow your own (GYO) kits labeled as containing G. lucidum were analyzed. DNA was extracted, and the internal transcribed spacer (ITS) region and translation elongation factor 1-alpha (tef1α) were sequenced with specific fungal primers. The majority (93%) of the manufactured reishi products and almost half of the GYO kits were identified as Ganoderma lingzhi. G. lingzhi is native to Asia and is the most widely cultivated and studied taxon for medicinal use. Illumina MiSeq sequencing of the ITS1 region was performed to determine if multiple Ganoderma species were present. None of the manufactured products tested contained G. lucidum sensu stricto, and it was detected in only one GYO kit. G. lingzhi was detected in most products, but other Ganoderma species were also present, including G. applanatum, G. australe, G. gibbosum, G. sessile, and G. sinense. Our results indicate that the content of these products vary and that better labeling is needed to inform consumers before these products are ingested or marketed as medicine. Of the 17 GYO kits tested, 11 kits contained Ganoderma taxa that are not native to the United States. If fruiting bodies of exotic Ganoderma taxa are cultivated, these GYO kits will likely end up in the environment. The effects of these exotic species to natural ecosystems needs investigation

Understudied, underrepresented, and unknown: Methodological biases that limit detection of early diverging fungi from environmental samples.

Metabarcoding is an important tool for understanding fungal communities. The internal transcribed spacer (ITS) rDNA is the accepted fungal barcode but has known problems. The large subunit (LSU) rDNA has also been used to investigate fungal communities but available LSU metabarcoding primers were mostly designed to target Dikarya (Ascomycota + Basidiomycota) with little attention to early diverging fungi (EDF). However, evidence from multiple studies suggests that EDF comprise a large portion of unknown diversity in community sampling. Here, we investigate how DNA marker choice and methodological biases impact recovery of EDF from environmental samples. We focused on one EDF lineage, Zoopagomycota, as an example. We evaluated three primer sets (ITS1F/ITS2, LROR/LR3, and LR3 paired with new primer LR22F) to amplify and sequence a Zoopagomycota mock community and a set of 146 environmental samples with Illumina MiSeq. We compared two taxonomy assignment methods and created an LSU reference database compatible with AMPtk software. The two taxonomy assignment methods recovered strikingly different communities of fungi and EDF. Target fragment length variation exacerbated PCR amplification biases and influenced downstream taxonomic assignments, but this effect was greater for EDF than Dikarya. To improve identification of LSU amplicons we performed phylogenetic reconstruction and illustrate the advantages of this critical tool for investigating identified and unidentified sequences. Our results suggest much of the EDF community may be missed or misidentified with "standard" metabarcoding approaches and modified techniques are needed to understand the role of these taxa in a broader ecological context

Woodpeckers can act as dispersal vectors for fungi, plants, and microorganisms

Bird-mediated dispersal is presumed to be important in the dissemination of many different types of organisms, but concrete evidence remains scarce. This is especially true for biota producing microscopic propagules. Tree-dwelling birds, such as woodpeckers, would seem to represent ideal dispersal vectors for organisms growing on standing tree trunks such as epiphytic lichens and fungi. Here, we utilize bird natural history collections as a novel source of data for studying dispersal ecology of plants, fungi, and microorganisms. We screened freshly preserved specimens of three Finnish woodpecker species for microscopic propagules. Samples were taken from bird feet, and chest and tail feathers. Propagules were extracted using a sonication-centrifugation protocol, and the material obtained was studied using light microscopy. Diverse biological material was recovered from all specimens of all bird species, from all positions sampled. Most abundant categories of discovered biological material included bryophyte fragments, fungal spores, and vegetative propagules of lichens. Also, freshwater diatoms, bryophyte spores, algal cells, testate amebae, rotifers, nematodes, pollen, and insect scales were identified. The method developed here is applicable to living specimens as well, making it a versatile tool for further research. Our findings highlight the potential of bird-mediated dispersal for diverse organisms and showcase the use of natural history collections in ecological research.Peer reviewe

Fungicide-Mediated Shifts in the Foliar Fungal Community of an Invasive Grass

Invasive plants, which cause substantial economic and ecological impacts, acquire both pathogens and beneficial microbes in their introduced ranges. Communities of fungal endophytes are known to mediate impacts of pathogens on plant fitness but few studies have examined the temporal dynamics of fungal communities on invasive plants. The annual grass Microstegium vimineum, an invader of forests and riparian areas throughout the eastern United States, experiences annual epidemics of disease caused by Bipolaris pathogens. Our objective was to characterize the dynamics of foliar fungal communities on M. vimineum over a growing season during a foliar disease epidemic. First, we asked how the fungal community in the phyllosphere changed over 2 months that corresponded with increasing disease severity. Second, we experimentally suppressed disease with fungicide in half of the plots and asked how the treatment affected fungal community diversity and composition. We found increasingly diverse foliar fungal communities and substantial changes in community composition between timepoints using high-throughput amplicon sequencing of the internal transcribed spacer 2 region. Monthly fungicide application caused shifts in fungal community composition relative to control samples. Fungicide application increased diversity at the late-season timepoint, suggesting that it suppressed dominant fungicide sensitive taxa and allowed other fungal taxa to flourish. These results raise new questions regarding the roles of putative endophytes found in the phyllosphere given the limited number of pathogens known to cause disease on M. vimineum in its invasive range

HID-1 is required for sensing CO₂ level in the pharynx.

<p>(<b>A</b>) One-day-old adult <i>hid-1(yg316)</i> and N2 worms were exposed to 5%, 10%, or 20% CO₂ balanced with 21% O₂ and N₂. The pumping rate was measured under a dissecting microscope while the animals were exposed to the different gas mixtures. A gas mixture of 21% O₂ and 79% N₂ was used as a normal air control. (<b>B</b>) The inhibition of the pumping rate of the pharynx after exposure to high CO₂ level in <i>hid-1(yg316)</i> allele mutants is significantly reduced. Similarly, the inhibition of the pumping rate of the pharynx after exposure to high CO₂ level is reduced in other <i>hid-1</i> allele mutants (<i>sa772</i> and <i>sa1058</i>). Transgenic expression of HID-1 fused to eGFP in the <i>sa722</i> or <i>yg316</i> background (<i>hid-1(sa722</i>);HID-1::GFP or <i>hid-1(yg316)</i>;HID-1::GFP) is sufficient to restore the effect of high CO₂ level on the pumping rate back to the wild-type phenotype. In all experiments <i>N</i>≥30 animals. Different groups were compared by one-way ANOVA followed by <i>t</i> test. ***<i>P</i><.001. Error bars indicate SEM.</p