Genome and physiology of the ascomycete filamentous fungus Xeromyces bisporus, the most xerophilic organism isolated to date.

Xeromyces bisporus can grow on sugary substrates down to 0.61, an extremely low water activity. Its genome size is approximately 22 Mb. Gene clusters encoding for secondary metabolites were conspicuously absent; secondary metabolites were not detected experimentally. Thus, in its 'dry' but nutrient-rich environment, X. bisporus appears to have relinquished abilities for combative interactions. Elements to sense/signal osmotic stress, e.g. HogA pathway, were present in X. bisporus. However, transcriptomes at optimal (∼ 0.89) versus low aw (0.68) revealed differential expression of only a few stress-related genes; among these, certain (not all) steps for glycerol synthesis were upregulated. Xeromyces bisporus increased glycerol production during hypo- and hyper-osmotic stress, and much of its wet weight comprised water and rinsable solutes; leaked solutes may form a protective slime. Xeromyces bisporus and other food-borne moulds increased membrane fatty acid saturation as water activity decreased. Such modifications did not appear to be transcriptionally regulated in X. bisporus; however, genes modulating sterols, phospholipids and the cell wall were differentially expressed. Xeromyces bisporus was previously proposed to be a 'chaophile', preferring solutes that disorder biomolecular structures. Both X. bisporus and the closely related xerophile, Xerochrysium xerophilum, with low membrane unsaturation indices, could represent a phylogenetic cluster of 'chaophiles'.

Peptaibol, secondary-metabolite, and hydrophobin pattern of commercial biocontrol agents formulated with species of the Trichoderma harzianum complex.

The production of bioactive polypeptides (peptaibiotics) in vivo is a sophisticated adaptation strategy of both mycoparasitic and saprotrophic Trichoderma species for colonizing and defending their natural habitats. This feature is of major practical importance, as the detection of peptaibiotics in plant-protective Trichoderma species, which are successfully used against economically relevant bacterial and fungal plant pathogens, certainly contributes to a better understanding of these complex antagonistic interactions. We analyzed five commercial biocontrol agents (BCAs), namely Canna(®) , Trichosan(®) , Vitalin(®) , Promot(®) WP, and TrichoMax(®) , formulated with recently described species of the Trichoderma harzianum complex, viz. T. afroharzianum, T. simmonsii, and T. guizhouense. By using the well-established, HPLC/MS-based peptaibiomics approach, it could unequivocally be demonstrated that all of these formulations contained new and recurrent peptaibols, i.e., peptaibiotics carrying an acetylated N-terminus, the C-terminus of which is reduced to a 1,2-amino alcohol. Their chain lengths, including the amino alcohol, were 11, 14, and 18 residues, respectively. Peptaibols were also to be the dominating secondary metabolites in plate cultures of the four strains obtained from four of the Trichoderma- based BCAs, contributing 95% of the UHPLC-UV/VIS peak areas and 99% of the total ion count MS peak area from solid media. Furthermore, species-specific hydrophobins, as well as non-peptaibiotic secondary metabolites, were detected, the latter being known for their antifungal, siderophore, or plant-growth-promoting activities. Notably, none of the isolates produced low-molecular weight mycotoxins.

Front line defenders of the ecological niche! Screening the structural diversity of peptaibiotics from saprotrophic and fungicolous Trichoderma/Hypocrea species.

Approximately 950 individual sequences of non-ribosomally biosynthesised peptides are produced by the genus Trichoderma/Hypocrea that belong to a perpetually growing class of mostly linear antibiotic oligopeptides, which are rich in the non-proteinogenic α-aminoisobutyric acid (Aib). Thus, they are comprehensively named peptaibiotics. Notably, peptaibiotics represent ca. 80 % of the total inventory of secondary metabolites currently known from Trichoderma/Hypocrea. Their unique membrane-modifying bioactivity results from amphipathicity and helicity, thus making them ideal candidates in assisting both colonisation and defence of the natural habitats by their fungal producers. Despite this, reports on the in vivo-detection of peptaibiotics have scarcely been published in the past. In order to evaluate the significance of peptaibiotic production for a broader range of potential producers, we screened nine specimens belonging to seven hitherto uninvestigated fungicolous or saprotrophic Trichoderma/Hypocrea species by liquid chromatography coupled to electrospray high resolution mass spectrometry. Sequences of peptaibiotics found were independently confirmed by analysing the peptaibiome of pure agar cultures obtained by single-ascospore isolation from the specimens. Of the nine species examined, five were screened positive for peptaibiotics. A total of 78 peptaibiotics were sequenced, 56 (=72 %) of which are new. Notably, dihydroxyphenylalaninol and O-prenylated tyrosinol, two C-terminal residues, which have not been reported for peptaibiotics before, were found as well as new and recurrent sequences carrying the recently described tyrosinol residue at their C-terminus. The majority of peptaibiotics sequenced are 18- or 19-residue peptaibols. Structural homologies with 'classical representatives' of subfamily 1 (SF1)-peptaibiotics argue for the formation of transmembrane ion channels, which are prone to facilitate the producer capture and defence of its substratum.

Phylogenetic relationships among members of the Fusarium solani species complex in human infections and the descriptions of F. keratoplasticum sp. nov. and F. petroliphilum stat. nov.

Fusarium species are frequently associated with mycotic keratitis and, to a lesser extent, cases of localized and disseminated infections. The Fusarium solani species complex (FSSC) is the most common group of fusaria associated with human infectious diseases. Several studies to date have revealed dozens of strongly supported phylogenetic species within this important evolutionary clade, though little work has been done to improve the taxonomy and understanding of the reproductive mode and phenotypes of the predominant clinically relevant species. Here we described Fusarium keratoplasticum sp. nov., and Fusarium petroliphilum stat. nov., two phylogenetic species that are among the most frequently isolated fusaria in plumbing drain biofilms and outbreaks of contact lens-associated mycotic keratitis. F. keratoplasticum isolates were highly variable and showed a range of morphological characteristics typical for most classical concepts of 'F. solani.' Many isolates failed to produce sporodochia and macroconidia. Although most attempts to sexually cross F. keratoplasticum isolates failed, a heterothallic sexual stage typical for the FSSC was discovered by pairing isolates of opposite mating type on V-8 agar, the ascospores of which showed molecular evidence of recombination. Secondary metabolite profiles of FSSC species defined through molecular data were compared for the first time and revealed the production of bioactive compounds including cyclosporines and several novel compounds of unknown function. We speculate that the inferred phenotypic variability in these species is the result of the almost entirely anthropogenic sources from which they are derived, including biofilms on plumbing systems.

Screening the biosphere: the fungicolous fungus Trichoderma phellinicola, a prolific source of hypophellins, new 17-, 18-, 19-, and 20-residue peptaibiotics.

To investigate the significance of antibiotics for the producing organism(s) in the natural habitat, we screened a specimen of the fungicolous fungus Trichoderma phellinicola (syn. Hypocrea phellinicola) growing on its natural host Phellinus ferruginosus. Results revealed that a particular group of non-ribosomal antibiotic polypeptides, peptaibiotics, which contain the non-proteinogenic marker amino acid, α-aminoisobutyric acid, was biosynthesized in the natural habitat by the fungicolous producer and, consequently, released into the host. By means of liquid chromatography coupled to electrospray high-resolution time-of-flight mass spectrometry, we detected ten 20-residue peptaibols in the specimen. Sequences of peptaibiotics found in vivo were independently confirmed by analyzing the peptaibiome of an agar plate culture of T. phellinicola CBS 119283 (ex-type) grown under laboratory conditions. Notably, this strain could be identified as a potent producer of 39 new 17-, 18-, and 19-residue peptaibiotics, which display the same building scheme as the 20-residue peptaibols found in the specimen. Two of the 19-residue peptaibols are tentatively assigned to carry tyrosinol, a novel C-terminal residue, as deduced from high-resolution tandem mass-spectrometry data. For the new peptaibiotics produced by T. phellinicola, the name 'hypophellin(s)', based on the teleomorph name, is introduced.

Production of fumonisins B2 and B4 in Tolypocladium species.

Tolypocladium inflatum is known primarily for its production of the cyclosporines that are used as an immunosuppressive drug. However, we report here the production of the carcinogenic fumonisins B(2) and B(4) by this biotechnologically relevant fungal genus. These mycotoxins were detected in 11 strains tested from three species: Tolypocladium inflatum, T. cylindrosporum, and T. geodes. Production of fumonisins by Fusarium spp. and Aspergillus niger is highly medium- and temperature-dependent, so the effect of these parameters on fumonisin production by three T. inflatum strains was studied. Maximum production was achieved on media with high sugar content incubated at 25-30°C. Since these results demonstrate that fumonisin production could be widespread within the genus Tolypocladium, the potential contamination of commercial cyclosporine preparations with fumonisins needs to be investigated.

A Pilot Study on Baseline Fungi and Moisture Indicator Fungi in Danish Homes.

In many complaint cases regarding bad indoor environments, there is no evidence of visible fungal growth. To determine if the problems are fungi-related, dust sampling is the method of choice among building surveyors. However, there is a need to differentiate between species belonging to a normal, dry indoor environment and species belonging to a damp building envelope. The purposes of this pilot study were to examine which fungal species are present in problem-free Danish homes and to evaluate different detection and identification methods. Analyses showed that the fungal diversity outside was different from the diversity inside and that the composition of fungal species growing indoors was different compared to those found as spores, both indoors and outdoors. Common for most homes were Pseudopithomyces chartarum, Cladosporium allicinum and Alternaria sect. Infectoriae together with Botrytis spp., Penicillium digitatum and Pen. glabrum. The results show that ITS sequencing of dust samples is adequate if supported by thorough building inspections and that food products play as large a role in the composition of the baseline spora as the outdoor air and surrounding vegetation. This pilot study provides a list of baseline fungal species found in Danish homes with a good indoor environment.

Fusarium spp. associated with rice Bakanae: ecology, genetic diversity, pathogenicity and toxigenicity.

African and Asian populations of Fusarium spp. (Gibberella fujikuroi species complex) associated with Bakanae of rice (Oryzae sativa L.) were isolated from seeds and characterized with respect to ecology, phylogenetics, pathogenicity and mycotoxin production. Independent of the origin, Fusarium spp. were detected in the different rice seed samples with infection rate ranges that varied from 0.25% to 9%. Four Fusaria (F. andiyazi, F. fujikuroi, F. proliferatum and F. verticillioides) were found associated with Bakanae of rice. While three of the Fusaria were found in both African and Asian seed samples, F. fujikuroi was only detected in seed samples from Asia. Phylogenetic studies showed a broad genetic variation among the strains that were distributed into four different genetic clades. Pathogenicity tests showed that all strains reduced seed germination and possessed varying ability to cause symptoms of Bakanae on rice, some species (i.e. F. fujikuroi) being more pathogenic than others. The ability to produce fumonisins (FB(1) and FB(2)) and gibberellin A3 in vitro also differed according to the Fusarium species. While fumonisins were produced by most of the strains of F. verticillioides and F. proliferatum, gibberellin A3 was only produced by F. fujikuroi. Neither fumonisin nor gibberellin was synthesized by most of the strains of F. andiyazi. These findings provide new information on the variation within the G. fujikuroi species complex associated with rice seed and Bakanae disease.

Effect of temperature and water activity on the production of fumonisins by Aspergillus niger and different Fusarium species.

BACKGROUND: Fumonisins are economically important mycotoxins which until recently were considered to originate from only a few Fusarium species. However recently a putative fumonisin gene cluster was discovered in two different Aspergillus niger strains followed by detection of an actual fumonisin B2 (FB2) production in four strains of this biotechnologically important workhorse. RESULTS: In the present study, a screening of 5 A. niger strains and 25 assumed fumonisin producing Fusarium strains from 6 species, showed that all 5 A. niger strains produced FB2 and 23 of 25 Fusarium produced fumonisin B1 and other isoforms (fumonisin B2 and B3). Five A. niger and five Fusarium spp. were incubated at six different temperatures from 15-42 degrees C on Czapek Yeast Agar +5% salt or Potato Dextrose Agar. A. niger had the highest production of FB2 at 25-30 degrees C whereas Fusarium spp. had the maximal production of FB1 and FB2 at 20-25 degrees C. Addition of 2.5-5% NaCl, or 10-20% sucrose increased the FB2 production of A. niger, whereas addition of glycerol reduced FB2 production. All three water activity lowering solutes reduced the fumonisin production of the Fusarium species. CONCLUSION: The present study shows that the regulation of fumonisin production is very different in A. niger and Fusarium, and that food and feeds preserved by addition of sugar or salts may be good substrates for fumonisin B2 production by A. niger.

Identification of potentially safe promising fungal cell factories for the production of polyketide natural food colorants using chemotaxonomic rationale.

BACKGROUND: Colorants derived from natural sources look set to overtake synthetic colorants in market value as manufacturers continue to meet the rising demand for clean label ingredients - particularly in food applications. Many ascomycetous fungi naturally synthesize and secrete pigments and thus provide readily available additional and/or alternative sources of natural colorants that are independent of agro-climatic conditions. With an appropriately selected fungus; using in particular chemotaxonomy as a guide, the fungal natural colorants could be produced in high yields by using the optimized cultivation technology. This approach could secure efficient production of pigments avoiding use of genetic manipulation. RESULTS: Polyketide pigment producing ascomycetous fungi were evaluated for their potential as production organisms based on a priori knowledge on species-specific pigment and potential mycotoxin production and BioSafety level (BSL) classification. Based on taxonomic knowledge, we pre-selected ascomycetous fungi belonging to Penicillium subgenus Biverticillium that produced yellow, orange or red pigments while deselecting Penicillium marneffei; a well known human pathogen in addition to other mycotoxigenic fungi belonging to the same group. We identified 10 strains belonging to 4 species; viz. P. purpurogenum, P. aculeatum, P. funiculosum, and P. pinophilum as potential pigment producers that produced Monascus-like pigments but no known mycotoxins. The selection/deselection protocol was illustrated in the pigment extracts of P. aculeatum IBT 14259 and P. crateriforme IBT 5015 analysed by HPLC-DAD-MS. In addition, extracellular pigment producing ability of some of the potential pigment producers was evaluated in liquid media with a solid support and N-glutarylmonascorubramine was discovered in the partially purified pigment extract of P. purpurogenum IBT 11181 and IBT 3645. CONCLUSION: The present work brought out that the use of chemotaxonomic tools and a priori knowledge of fungal extrolites is a rational approach towards selection of fungal polyketide pigment producers considering the enormous chemical diversity and biodiversity of ascomycetous fungi. This rationale could be very handy for the selection of potentially safe fungal cell factories not only for polyketide pigments but also for the other industrially important polyketides; the molecular and genetic basis for the biosynthesis of which has not yet been examined in detail. In addition, 4 out of the 10 chemotaxonomically selected promising Penicillium strains were shown to produce extracellular pigments in the liquid media using a solid support indicating future cell factory possibilities for polyketide natural food colorants.

Evaluation of Epicoccum nigrum for growth, morphology and production of natural colorants in liquid media and on a solid rice medium.

Four nonpathogenic and nontoxigenic Epicoccum nigrum strains were evaluated for their growth, morphology and pigment producing ability in three complex and one defined liquid media. Epicoccum nigrum IBT 41028 produced pigments in all the four media tested with a maximum pigment of 3.68 AU at 410 nm in M1 medium (unoptimized) containing 5 g/l yeast autolysate. The color hue of the crude pigment extracts ranged from 74 to 102 exhibiting dark orange to green-yellow color. Pelleted morphology was shown to have a positive influence on the pigment production by E. nigrum strain IBT 41028 in the liquid media, and the use of Bis-tris buffer was found to diminish or reduce the pellet formation. Since Monascus is a well known pigment producer on rice. Pigment producing ability of E. nigrum IBT 41028 was tested on rice and compared to liquid media with Monascus ruber IBT 7904 as control. Though, both genera preferred rice but E. nigrum produced 4.6 folds higher pigment in the liquid unoptimized fermentation medium compared to M. ruber. Solid phase extraction and subsequently HPLC-DAD analysis of the crude pigment extracts showed qualitative as well as quantitative variation in the pigment composition under solid and liquid cultivations.

Heterologous production of the widely used natural food colorant carminic acid in Aspergillus nidulans.

The natural red food colorants carmine (E120) and carminic acid are currently produced from scale insects. The access to raw material is limited and current production is sensitive to fluctuation in weather conditions. A cheaper and more stable supply is therefore desirable. Here we present the first proof-of-concept of heterologous microbial production of carminic acid in Aspergillus nidulans by developing a semi-natural biosynthetic pathway. Formation of the tricyclic core of carminic acid is achieved via a two-step process wherein a plant type III polyketide synthase (PKS) forms a non-reduced linear octaketide, which subsequently is folded into the desired flavokermesic acid anthrone (FKA) structure by a cyclase and a aromatase from a bacterial type II PKS system. The formed FKA is oxidized to flavokermesic acid and kermesic acid, catalyzed by endogenous A. nidulans monooxygenases, and further converted to dcII and carminic acid by the Dactylopius coccus C-glucosyltransferase DcUGT2. The establishment of a functional biosynthetic carminic acid pathway in A. nidulans serves as an important step towards industrial-scale production of carminic acid via liquid-state fermentation using a microbial cell factory.

On the biosynthetic origin of carminic acid.

The chemical composition of the scale insect Dactylopius coccus was analyzed with the aim to discover new possible intermediates in the biosynthesis of carminic acid. UPLC-DAD/HRMS analyses of fresh and dried insects resulted in the identification of three novel carminic acid analogues and the verification of several previously described intermediates. Structural elucidation revealed that the three novel compounds were desoxyerythrolaccin-O-glucosyl (DE-O-Glcp), 5,6-didehydroxyerythrolaccin 3-O-ß-D-glucopyranoside (DDE-3-O-Glcp), and flavokermesic acid anthrone (FKA). The finding of FKA in D. coccus provides solid evidence of a polyketide, rather than a shikimate, origin of coccid pigments. Based on the newly identified compounds, we present a detailed biosynthetic scheme that accounts for the formation of carminic acid (CA) in D. coccus and all described coccid pigments which share a flavokermesic acid (FK) core. Detection of coccid pigment intermediates in members of the Planococcus (mealybugs) and Pseudaulacaspis genera shows that the ability to form these pigments is taxonomically more widely spread than previously documented. The shared core-FK-biosynthetic pathway and wider taxonomic distribution suggests a common evolutionary origin for the trait in all coccid dye producing insect species.

Analysis of moniliformin in maize plants using hydrophilic interaction chromatography.

A novel HPLC method was developed for detection of the Fusarium mycotoxin, moniliformin in whole maize plants. The method is based on hydrophilic interaction chromatography (HILIC) on a ZIC zwitterion column combined with diode array detection and negative electrospray mass spectrometry (ESI(-)-MS). Samples were extracted using acetonitrile-water (85:15), and the extracts were cleaned up on strong anion exchange columns. By this procedure we obtained a recovery rate of 57-74% moniliformin with a limit of detection at 48 ng/g and a limit of quantification at 96 ng/g using UV detection at 229 nm, which is comparable to current methods used. Limit of detection and quantification using ESI(-)-MS detection was 1 and 12 ng/g, respectively. Screening of maize samples infected with the moniliformin producing fungi F. avenaceum, F. tricinctum, or F. subglutinans detected moniliformin levels of 1-12 ng/g in 15 of 28 samples using ESI(-)-MS detection. To our knowledge this is the first example of HILIC separation in mycotoxin analysis.

Fumonisin B2 production by Aspergillus niger.

The carcinogenic mycotoxin fumonisin B2 was detected for the first time in the industrially important Aspergillus niger. Fumonisin B2, known from Fusarium verticillioides and other Fusaria, was detected in cultures of three full genome sequenced strains of A. niger, in the ex type culture and in a culture of F. verticillioides by electrospray LC-MS analysis of methanolic extracts from agar plugs of cultures grown on several substrates. Whereas F. verticillioides produced fumonisins B1, B2, and B3 on agar media based on plant extracts, such as barley malt, oat, rice, potatoes, and carrots, A. niger produced fumonisin B2 best on agar media with a low water activity, including Czapek yeast autolysate agar with 5% NaCl. Of the media tested, only rice corn steep agar supported fumonisin production by both F. verticillioides and A. niger. However, A. niger had a different regulation of fumonisin production and a different quantitative profile of fumonisins, producing only B2 as compared to F. verticillioides. Fumonisin production by A. niger, which is a widely occurring species and an extremely important industrial organism, will have very important implications for biotechnology and especially food safety. A. niger is used for the production of citric acid and as producer of extracellular enzymes, and also as a transformation host for the expression of heterologous proteins. Certain strains of A. niger produce both ochratoxin A and fumonisins, so some foods and feeds may potentially contain two types of carcinogenic mycotoxins from this species.

Genome Sequence of Talaromyces atroroseus, Which Produces Red Colorants for the Food Industry.

Talaromyces atroroseus is a known producer of Monascus colorants suitable for the food industry. Furthermore, genetic tools have been established that facilitate elucidation and engineering of its biosynthetic pathways. Here, we report the draft genome of a potential fungal cell factory, T. atroroseus IBT 11181 (CBS 123796).

Genes Linked to Production of Secondary Metabolites in Talaromyces atroroseus Revealed Using CRISPR-Cas9.

The full potential of fungal secondary metabolism has until recently been impeded by the lack of universal genetic tools for most species. However, the emergence of several CRISPR-Cas9-based genome editing systems adapted for several genera of filamentous fungi have now opened the doors for future efforts in discovery of novel natural products and elucidation and engineering of their biosynthetic pathways in fungi where no genetic tools are in place. So far, most studies have focused on demonstrating the performance of CRISPR-Cas9 in various fungal model species, and recently we presented a versatile CRISPR-Cas9 system that can be successfully applied in several diverse Aspergillus species. Here we take it one step further and show that our system can be used also in a phylogenetically distinct and largely unexplored species from the genus of Talaromyces. Specifically, we exploit CRISPR-Cas9-based genome editing to identify a new gene in T. atroroseus responsible for production of polyketide-nonribosomal peptide hybrid products, hence, linking fungal secondary metabolites to their genetic origin in a species where no genetic engineering has previously been performed.

Exploring fungal biodiversity for the production of water-soluble pigments as potential natural food colorants.

The production of many currently authorized natural food colorants has a number of disadvantages, including a dependence on the supply of raw materials and variations in pigment extraction. Fungi provide a readily available alternative source of naturally derived food colorants that could easily be produced in high yields. The recent authorization of a fungal food colorant has fuelled research to explore the extraordinary chemical diversity and biodiversity of fungi for the biotechnological production of pigments as natural food colorants. These studies require an appropriate use of chemotaxonomic tools and a priori knowledge of fungal metabolites to carry out intelligent screening for known or novel colorants as lead compounds. Such screening would result in the preselection of some potential pigment producers and the deselection of pathogenic strains and toxin producers. With advances in gene technology, in the future it should be possible to employ metabolic engineering to create microbial cell factories for the production of food colorants.

Colorimetric characterization for comparative analysis of fungal pigments and natural food colorants.

Exogenous pigments produced by ascomycetous filamentous fungi belonging to the genera Penicillium, Epicoccum, and Monascus, preselected based on chemotaxonomic knowledge, have been extracted and characterized by quantitative colorimetry. The color characteristics of the fungal extracts were compared to water soluble natural colorants derived from sources currently in use. The tested fungal extracts also included some commercially available Monascus colorants. The a values for the fungal extracts were found to be both positive and negative, the b values were found to be positive, while the hue angles of the fungal color extracts ranged from 40 to 110 indicating the color distribution of fungal extracts over the red-orange-yellow region of the CIELAB color space. The fungal extracts exhibited additional color hues in the red spectrum and similar hues in the yellow spectrum as compared to the reference natural colorants. They were also found to be similar or brighter in terms of chroma to some of the reference natural colorants. Principal component analysis was performed to group and distinguish different colors based on the a and b values. The fungal color extracts could be grouped in accordance with the similarity or difference in the color to those of the existing natural colorants. The diversity of colors was not only found among different fungal genera and/or species but also within the same species on changing the media. There was a marked change in the color composition of the extracts resulting in relatively different hues. Our results, thus, indicate that there exists pigment-producing genera of ascomycetous fungi other than Monascus that produce color shades in the red and the yellow spectra in addition or similar to reference colorants. These color shades could add to the color palette of the natural colorants currently in use. In addition, the multivariate approach in distinguishing and classifying the colorants was shown to be a very useful tool in colorimetric comparison of colorants.

Draft genome sequence and chemical profiling of Fusarium langsethiae, an emerging producer of type A trichothecenes.

Fusarium langsethiae is a widespread pathogen of small grain cereals, causing problems with T-2 and HT-2 toxin contamination in grains every year. In an effort to better understand the biology of this fungus, we present a draft genome sequence of F. langsethiae Fl201059 isolated from oats in Norway. The assembly was fragmented, but reveals a genome of approximately 37.5 Mb, with a GC content around 48%, and 12,232 predicted protein-coding genes. Focusing on secondary metabolism we identified candidate genes for 12 polyketide synthases, 13 non-ribosomal peptide synthetases, and 22 genes for terpene/isoprenoid biosynthesis. Some of these were found to be unique compared to sequence databases. The identified putative Tri5 cluster was highly syntenic to the cluster reported in F. sporotrichioides. Fusarium langsethiae Fl201059 produces a high number of secondary metabolites on Yeast Extract Sucrose (YES) agar medium, dominated by type A trichothecenes. Interestingly we found production of glucosylated HT-2 toxin (Glu-HT-2), previously suggested to be formed by the host plant and not by the fungus itself. In greenhouse inoculations of F. langsethiae Fl201059 on barley and oats, we detected the type A trichothecenes: neosolaniol, HT-2 toxin, T-2 toxin, Glu-HT-2 and numerous derivatives of these.