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Many species of fungi have been detected in the healthy human gut; however, nearly half of all taxa reported have only been found in one sample or one study. Fungi capable of growing in and colonizing the gut are limited to a small number of species, mostlyCandidayeasts and yeasts in the family Dipodascaceae (Galactomyces, Geotrichum, Saprochaete).Malasseziaand the filamentous fungusCladosporiumare potential colonizers; more work is needed to clarify their role. Other commonly-detected fungi come from the diet or environment but either cannot or do not colonize (PenicilliumandDebaryomycesspecies, which are common on fermented foods but cannot grow at human body temperature), while still others have dietary or environmental sources (Saccharomyces cerevisiae, a fermentation agent and sometime probiotic;Aspergillusspecies, ubiquitous molds) yet are likely to impact gut ecology. The gut mycobiome appears less stable than the bacterial microbiome, and is likely subject to environmental factors. [ABSTRACT FROM PUBLISHER]

We have entered the Age of the Microbiome, with new studies appearing constantly and whole journals devoted to the human microbiome. While bacteria outnumber other gut microbes by orders of magnitude, eukaryotes are consistently found in the human gut and are represented primarily by the fungi. Compiling 36 studies 1917-2015 we found at least 267 distinct fungal taxa have been reported from the human gut, and seemingly every new study includes one or more fungi not previously described from this niche. This diversity, while impressive, is illusory. If we examine gut fungi, we will quickly observe a division between a small number of commonly detected species (Candida yeasts, Saccharomyces and yeasts in the Dipodascaceae, and Malassezia species) and a long tail of taxa that have been reported only once. Furthermore, an investigation into the ecology of these rare species reveals that many of them are incapable of colonization or long-term persistence in the gut. This paper examines what we know and have yet to learn about the fungal component of the gut microbiome, or "mycobiome", and an overview of methods. We address the potential of the field while introducing some caveats and argue for the necessity of including mycologists in mycobiome studies. [ABSTRACT FROM AUTHOR]

This experiment exposed 3rd and 4th y undergraduates and graduate students taking a course in advanced food analysis to DNA extraction, polymerase chain reaction (PCR), and DNA sequence analysis. Students provided their own fish sample, purchased from local grocery stores, and the class as a whole extracted DNA, which was then subjected to PCR, gel electrophoresis, and Sanger sequencing. Students retrieved their sequences and identified their fish using the NCBI BLAST nucleotide database. Slightly more than half of the samples yielded sequences identical or close to expected (based on the identification of the fish on the packaging); some other samples matched unanticipated fish or other organisms, due to an incomplete database, minor sequencing errors, or laboratory contamination (human and fungal sequences); 1 canned tuna sample identified as hake could represent food fraud. [ABSTRACT FROM AUTHOR]

Abstract: 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 Galerina marginata, 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 Galerina hybrida 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. [Copyright &y& Elsevier]

Deoxynivalenol (DON) is a potent mycotoxin and virulence factor produced by Fusarium graminearum. We examined the expression of the core DON biosynthetic gene Tri5 during wheat head infection of susceptible and resistant cultivars and susceptible cultivars treated with strobilurin fungicides (e.g., azoxystrobin). DON was quantified to correlate expression with toxin accumulation. The highest Tri5 expression relative to housekeeping genes occurred at the infection front. As infection progressed, earliest-infected kernels showed diminished relative Tri5 expression but Tri5 expression never ceased during the 21 days observed. Azoxystrobin treatment showed no significant effect on either relative Tri5 expression or DON quantity. The resistant cultivar 'Alsen' showed minimal spread of the fungus, with no fungus detected by day 21. DON was not detected in significant quantities in Alsen in the later stages sampled. In Wheaten. DON levels were negligible at 8 days post-inoculation (dpi), with detectable DON at later-sampled lime points. Tri5 was detected even in fully senesced kernels 21 dpi. Our data demonstrate the presence of Tri5 transcripts in a susceptible cultivar over a much longer time period than has been previously documented. This suggests the ability of the fungus to rapidly resume toxin biosynthesis in dried infected grain should conducive environmental conditions be present, and provides a possible mechanism for high DON levels in asymptomatic grain. [ABSTRACT FROM AUTHOR]

The peptide toxins of poisonous Amanita mushrooms are bicyclic octapeptides (amatoxins) or heptapeptides (phallotox ins). In Amanita bisporigera, α-amanitin and phallacidin are synthesized as 35- and 34-amino acid proproteins, respectively, in which the amino acid sequences found in the mature toxins are flanked by conserved amino acid sequences. The presence of invariant Pro residues immediately upstream of the toxin regions and as the last predicted amino acid in the toxin regions themselves suggests that a Prospecific peptidase is responsible for the initial posttranslational processing of the Amanita toxin proproteins. We purified an enzyme from the phalloidin producing mushroom Conocybe albipes that cleaves a synthetic 22-mer phalloidin peptide to release the mature toxin peptide (AWLATCP). Mass spectrometric analysis of the purified pro tein combined with isolation and sequencing of the encoding gene indicates that the responsible processing enzyme is a member of the prolyl oligopeptidase (POP) subfamily of proteases (EC 3.4.21.26). The processing enzyme was able to use the chromogenic POP substrate benzyloxycarbonyl-Gly-Pro-p-nitro-anilide and was inhibited by the specific POP inhibitor benzyl oxycarbonyl-Pro-prolinal. Both Pro bonds in the pro-protein are cleaved by the same enzyme, with the Cterminal Pro bond cleaved first or much faster than the N-terminal Pro bond. Transient accumulation of the N-terminal intermediate indicates that cleavage is not strongly processive. A synthetic peptide rep resenting the phallacidin pro-protein was also cleaved by the POP of C. albipes, but a precursor of amanitin (which is not made by C. albipes) was cleaved inefficiently. [ABSTRACT FROM AUTHOR]

Abstract: Based on the analysis of its genome sequence, the ectomycorrhizal (ECM) basidiomycetous fungus Laccaria bicolor was shown to be lacking many of the major classes of secreted enzymes that depolymerize plant cell wall polysaccharides. To test whether this is also a feature of other ECM fungi, we searched a survey genome database of Amanita bisporigera with the proteins found in the secretome of Trichoderma reesei (syn. Hypocrea jecorina), a biochemically well-characterized industrial fungus. Additional proteins were also used as queries to compensate for major groups of cell-wall-degrading enzymes lacking in the secretome of T. reesei and to substantiate conclusions drawn from the T. reesei collection. By MS/MS-based “shotgun” proteomics, 80 proteins were identified in culture filtrates of T. reesei strain RUTC30 grown on corn cell walls and in a commercial “cellulase” preparation, Spezyme CP. The two T. reesei enzyme preparations were qualitatively and quantitatively similar, the most striking difference being the lack of at least five major peptidases from the commercial enzyme mixture. Based on our analysis of A. bisporigera, this ECM fungus is deficient in many major classes of cell-wall-degrading enzymes, including both glycosyl hydrolases and carbohydrate esterases. By comparison, the genomes of the saprophytic basidiomycetes Coprinopsis cinerea and Galerina marginata (using a genome survey sequence approximately equivalent in depth to that of A. bisporigera) have, like T. reesei, a much more complete complement of cell-wall-degrading enzymes. [Copyright &y& Elsevier]

The systematic position of secotioid (Torrendia) and gasteroid (Amarrendia) forms within the agaricoid Amanita lineage (Agaricales, Basidiomycota) was studied with molecular (nLSU, ITS) data. Secotioid and gasteroid forms occur in fore" independent clades nested within agaricoid forms. One clade corresponds to the secotioid T. pulchella from southern Europe and northern Africa. The others correspond to Torrendia and Amarrendia species from Australia. Mediterranean climatic conditions are postulated as a force driving the convergent evolution of these secotioid and at least one of the gasteroid forms in geographically distant areas. Species formerly placed in Torrendia and Amarrendia are transferred to Amanita. A new species of Torrrendia from Australia was discovered during the revision of the collections originally identified as T. arenaria and is described here as Amanita pseudoinculta. [ABSTRACT FROM AUTHOR]

Fusarium head blight (FHB) and the associated mycotoxin deoxynivalenol (DON) negatively impact the wheat industry worldwide. In North America, FHB is mainly caused by Fusarium graminearum sensu stricto. The purpose of this study was to evaluate, under storage conditions, the expression of the DON biosynthetic gene trichodiene synthase (Tri5) of F. graminearum in grain of hard red winter wheat cultivars 'Overley' (FHB-susceptible) and 'Overland' (moderately resistant to FHB) from field plots treated or untreated with the triazole fungicide Prosaro and the strobilurin fungicide Headline. Infected grain was stored and periodically sampled to determine gene expression by qRT-PCR analysis. The F. graminearum housekeeping gene GAPDH was consistently detected, indicative of metabolically active fungi, and Tri5 detection was significantly higher in 'Overley' compared with 'Overland'. Tri5 gene expression and DON concentrations showed little to no correlation; consequently, Tri5 expression levels did not accurately predict DON concentrations. The strobilurin did not significantly reduce Tri5 gene expression compared with untreated wheat. In the triazole treatment, a significant reduction in the relative expression of Tri5 was detected after 120 days, as well as a downregulation of Tri5 from 60 to 120 days of storage in 'Overley'. In grain from strobilurin-treated plots of both cultivars, the expression of Tri5 increased from 0 to 30 days after tempering. Genetic expression of Tri5 that is necessary for the production of DON can increase during storage of high-moisture grain. Fusarium fungi can persist in wheat kernels for several months postharvest and may actively produce toxin during this period. [ABSTRACT FROM AUTHOR]

Several Fusarium species cause Fusarium head blight (FHB), a devastating disease that affects wheat and other small grain cereals. Besides causing significant yield losses, FHB pathogens reduce the quality of the grain and may produce mycotoxins that are harmful to both humans and animals. The development of effective management strategies for this disease requires an understanding of the composition of the FHB pathogen populations, which is important because different populations could be affected selectively. Recent reports of shifts in populations of FHB pathogens have shown that these populations are dynamic and change continuously, often associated with increased yield losses or changes in the mycotoxins produced in the grain. These population shifts include: replacement of F. graminearum by F. poae in Italy, hypothesized to be due to variation in environmental conditions; replacement of F. culmorum by F. graminearum in Europe, associated with climate change and increased maize production; a more aggressive and toxigenic F. asiaticum 3-ADON population replacing the existing NIV population in China; and a highly toxigenic population composed mainly of F. graminearum 3-ADON isolates replacing the existing 15-ADON population in North America. Additionally, reports of recent introductions of foreign species or populations into new areas include: F. asiaticum outside of Asia, F. graminearum NIV isolates in the USA and Luxembourg, F. graminearum 15-ADON isolates in Norway, F. graminearum in South Korea, F. boothii in Europe and USA, F. vorosii in Hungary and F. cortaderiae in New Zealand and Europe. [ABSTRACT FROM AUTHOR]

The biosynthetic pathway for amanitins and related cyclic peptides in deadly Amanita (Amanitaceae) mushrooms represents the first known ribosomal cyclic peptide pathway in the Fungi. Amanitins are found outside of the genus in distantly related agarics Galerina (Strophariaceae) and Lepiota (Agaricaceae). A long-standing question in the field persists: why is this pathway present in these phylogenetically disjunct agarics? Two deadly mushrooms, A. pallidorosea and A. subjunquillea, were deep sequenced, and sequences of biosynthetic genes encoding MSDINs (cyclic peptide precursor) and prolyl oligopeptidases (POPA and POPB) were obtained. The two Amanita species yielded 29 and 18 MSDINs, respectively. In addition, two MSDIN sequences were cloned from L. brunneoincarnata basidiomes. The toxin MSDIN genes encoding amatoxins or phallotoxins from the three genera were compared, and a phylogenetic tree constructed. Prolyl oligopeptidase B (POPB), a key enzyme in the biosynthetic pathway, was used in phylogenetic reconstruction to infer the evolutionary history of the genes. Phylogenies of POPB and POPA based on both coding and amino acid sequences showed very different results: while POPA genes clearly reflected the phylogeny of the host species, POPB did not; strikingly, it formed a well-supported monophyletic clade, despite that the species belong to different genera in disjunct families. POPA, a known house-keeping gene, was shown to be restricted in a branch containing only Amanita species and the phylogeny resembled that of those Amanita species. Phylogenetic analyses of MSDIN and POPB genes showed tight coordination and disjunct distribution. A POPB gene tree was compared with a corresponding species tree, and distances and substitution rates were compared. The result suggested POPB genes have significant smaller distances and rates than the house-keeping rpb2, discounting massive gene loss. Under this assumption, the incongruency between the gene tree and species tree was shown with strong support. Additionally, k-mer analyses consistently cluster Galerina and Amanita POPB genes, while Lepiota POPB is distinct. Our result suggests that horizontal gene transfer (HGT), at least between Amanita and Galerina, was involved in the acquisition of POPB genes, which may shed light on the evolution of the α-amanitin biosynthetic pathway. [ABSTRACT FROM AUTHOR]

Maize ( Zea mays) is a staple in many developing countries but is known to be prone to pest (insects, birds, and rodents) and fungal infestation. In Guatemala, mycotoxin contamination of cultivated products may occur owing to such factors as environmental conditions and the use of traditional agriculture operations. To assess the current maize conditions in Guatemala, a small-scale study was performed. Mold and insect counts and mycotoxin (aflatoxin and fumonisin) concentrations were determined on 25 farms in two townships (Chiantla and Todos Santos) of the Huehuetenango Department. Total fungal counts were 3.6 to 6.83 log CFU/g with no significant differences ( P > 0.05) across farms at different altitudes. Farms where maize was not produced but was purchased were at higher risk of fumonisin contamination, whereas local producers were mostly affected by aflatoxins. Aflatoxin was present in maize from 100% of farms at 1.0 to 85.3 ppb, and fumonisin was detected on 52% of farms at 0.4 to 31.0 ppm. Average mycotoxin consumption amounts were above the recommended maximum intake for aflatoxin in both produced and purchased maize and above the provisional maximum tolerable daily intake for fumonisin in purchased maize. Estimated daily intake was 0.01 to 0.85 μg/kg of body weight per day for aflatoxin and 2.9 to 310.0 μg/kg of body weight per day for fumonisin. An entomological analysis revealed overall 32% prevalence of Ephestia kuehniella (flour moth), 16% prevalence of Sitophilus zeamais (maize weevil), and 8% prevalence of Tribolium sp. (flour beetle) on the analyzed farms. This study highlighted poor agricultural practices used in the highlands of Guatemala. Current practices should be revised for the production of maize that is safe for consumption by the population in this region. [ABSTRACT FROM AUTHOR]

Small bowel transplantation (SBT) can be a life-saving medical procedure. However, these recipients experience high risk of bloodstream infections caused by Candida. This research aims to characterise the SBT recipient gut microbiota over time following transplantation and investigate the epidemiology of candidaemia in seven paediatric patients. Candida species from the recipients' ileum and bloodstream were identified by internal transcribed spacer sequence and distinguished to strain by multilocus sequence typing and randomly amplified polymorphic DNA. Antifungal susceptibility of bloodstream isolates was determined against nine antifungals. Twenty-two ileostomy samples harboured at least one Candida species. Fungaemia were caused by Candida parapsilosis, Candida albicans, Candida glabrata, Candida orthopsilosis and Candida pelliculosa. All but three bloodstream isolates showed susceptibility to all the antifungals tested. One C. glabrata isolate showed multidrug resistance to itraconazole, amphotericin B and posaconazole and intermediate resistance to caspofungin. Results are congruent with both endogenous ( C. albicans, C. glabrata) and exogenous ( C. parapsilosis) infections; results also suggest two patients were infected by the same strain of C. parapsilosis. Continuing to work towards a better understanding of sources of infection-particularly the exogenous sources-would lead to targeted prevention strategies. [ABSTRACT FROM AUTHOR]

Candida yeasts are the dominant fungi in the healthy human microbiome, but are well-known for causing disease following a variety of perturbations. Evaluation of fungal populations from the healthy human gut revealed a significant negative correlation between the foodborne yeast, Debaryomyces hansenii , and Candida species. D. hansenii is reported to produce killer toxins (mycocins) effective against other yeast species. In order to better understand this phenomenon, a collection of 42 D. hansenii isolates was obtained from 22 cheeses and evaluated for killer activity against Candida albicans and Candida tropicalis over a range of temperatures and pH values. Twenty three strains demonstrated killer activity against both C. albicans and C. tropicalis , which was pH- and temperature-dependent, with no killer activity observed for any strain at pH 6.5 or higher, or at ≥ 35 °C (physiological conditions in the human gastrointestinal tract). A cell-free mycocin preparation showed transient killer activity against C. albicans at 35 °C and a cheese sample containing a killer D. hansenii strain demonstrated sustained killer activity against both C. albicans and C. tropicalis . Together, these observations raise the possibility that D. hansenii could influence Candida populations in the gut. [ABSTRACT FROM AUTHOR]

Fusarium head blight (FHB) is a devastating disease of wheat (Triticum aestivum L.) and other small grain cereals. The disease is caused by several species of Fusarium. This study was conducted to identify the major species of Fusarium causing FHB in Nebraska and to characterize isolates of the species for perithecia and deoxynivalenol (DON) production and for aggressiveness (quantified as disease severity and area under the disease progress curve) on spikes of two winter wheat cultivars. Isolates of Fusarium spp. obtained from wheat spikes and grain collected from FHB-affected winter wheat fields and grain elevators, respectively, in 2007 and 2008 were identified as Fusarium graminearum. The isolates varied widely in perithecia and DON production in vitro and in aggressiveness on wheat spikes of soft red winter wheat cultivars Coker 9835 (FHB susceptible) and VA04W-433 (moderately resistant). Fusarium head blight severity for the three most aggressive F. graminearum isolates was higher in Coker 9835 than in VA04W-433 during the first 7 d after inoculation (DAI) but was comparable in both cultivars from 10 to 21 DAI. Deoxynivalenol concentration and aggressiveness were positively and linearly related (R² ≥ 0.60, P < 0.05). Isolates that produced higher concentrations of DON were more aggressive than those that produced lower concentrations of the toxin regardless of wheat cultivar. [ABSTRACT FROM AUTHOR]