Fusarium og mykotoksiner

Aksfusariose er en utbredt og destruktiv soppsykdom i korn forårsaket av en rekke arter innen soppslektene Fusarium og Microdochium. Aksfusariose kan redusere kornkvaliteten da ulike Fusarium-arter kan produsere en rekke giftige metabolitter (mykotoksiner) som kan være helseskadelige for mennesker og dyr. Foto: Jafar Razzaghian, NIBIO

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Foto: Jafar Razzaghian / NIBIO
KONTAKTPERSON
Medarbeidere
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Foto: Erling Fløistad

Fusarium og mykotoksiner i norsk korn

NIBIO har i ulike prosjekter kartlagt forekomst av Fusarium og mykotoksiner i norsk korn. Vi bruker både morfologiske og molekylære metoder for å identifisere og kvantifisere ulike sopparter. Kjemiske analysemetoder (LC-HRMS) brukes for å kvantifisere ulike mykotoksiner i korn ned til et nivå på 1 µg/kg.

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Foto: Jafar Razzaghian

Fusarium graminearum

Som i flere andre europeiske land har den relative utbredelsen av sopparter som produserer mykotoksinet deoksynivalenol (DON) endret seg fra Fusarium culmorum til F. graminearum. Fusarium graminearum er nå regnet som hovedprodusent av DON i norsk korn. NIBIO har kartlagt genetisk og fenotypisk variasjon innen norske isolater av F. graminearum

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Foto: Jafar Razzaghian

Effekt av værforhold på utvikling av Fusarium

Nedbør og temperatur i den kritiske perioden rundt blomstring har stor innvirkning på utviklingen av aksfusariose og mykotoksiner i korn. NIBIO arbeider med å utvikle prognosemodeller for å forutsi risiko for utvikling av mykotoksiner i korn på bakgrunn av dyrkningspraksis og værforhold.

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Foto: Till Seehusen

Effekt av dyrkingspraksis på utvikling av Fusarium 

Dyp jordbearbeiding og vekstskifte anses å være av stor betydning for bekjempelse av Fusarium og mykotoksiner i korn. NIBIO kartlegger betydningen av ulike tiltak som sortsvalg, jordarbeiding, forgrøde, jordart og fungicidbehandling på utvikling av Fusarium og mykotoksiner i korn.

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Foto: Ingerd Skow Hofgaard

Plante-patogen interaksjoner

Konkurranseevnen til ulike Fusarium-arter innen aksfusariose komplekset er påvirket av værforhold, dyrkingspraksis og vertsplante. Vi gjennomfører forsøk i felt, i drivhus- og på laboratoriet for å studere interaksjoner mellom ulike kornarter og sopper innen Fusarium spp. og Microdochium spp.

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Foto: Lars Sandved Dalen

Genomsekvensering av Fusarium

Kunnskap om gener involvert i plante-patogen interaksjoner kan være nyttig når man utvikler fremtidige bekjempelsesstrategier. For å kunne studere genuttrykk (transkriptom) av Fusarium under en infeksjon, har vi sekvensert genomene til F. langsethiae og F. avenaceum.

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Foto: Ingerd Skow Hofgaard

Resistens mot Fusarium i norsk korn

Ingen kommersielle sorter av hvete, havre eller bygg er absolutt resistente mot Fusarium spp., men det er funnet forskjeller i graden av resistens mellom ulike sorter. NIBIO gjennomfører feltforsøk, samt forsøk i drivhus- og i lab for å kartlegge resistens mot Fusarium spp. i ulike kornsorter. Ved å dyrke sorter med størst mulig grad av resistens mot Fusarium vil risikoen for utvikling av mykotoksiner i korn reduseres. 

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Foto: Erling Fløistad

Kjemisk og biologisk bekjempelse av aksfusariose

NIBIO undersøker effekten av biopestisider og fungicider på utvikling av Microdochium spp., Fusarium spp. og mykotoksiner i korn. Protiokonazol (det aktive stoffet i blant annet Proline) har vist seg å halvere DON-innholdet i havre sammenlignet med ubehandlet kontroll. Imidlertid har ingen av de testede biopestisidene resultert i en reduksjon av DON, og ingen fungicider har så langt vist seg å kunne begrense Fusarium langsethiae og HT2 + T2-toksiner i havre.

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Foto: Ingerd Skow Hofgaard

Fusarium og Microdochium på korn

Angrep av Fusarium / Microdochium kan føre til redusert kornkvalitet. Gjennom laboratorie-, drivhus- og feltforsøk undersøker NIBIO forekomst og overlevelse av disse patogenene i korn. Ofte i samarbeid med Kimen såvarelaboratoriet. Nylig har NIBIO i samarbeid med Nofima og NMBU (Norges miljø- og biovitenskapelige universitet) identifisert en mulig sammenheng mellom forekomst av Fusarium / Microdochium spp. og bakekvalitet i hvete.

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Fra bukkefôr og husmannskost til urban hipstermat; havre har vært en viktig komponent i kosthold og fôr i generasjoner. Men noen havresorter er mer sårbare for soppgifter enn andre. Dyrking av mer resistente sorter kan redusere risikoen for soppgifter i mat og fôr.

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For bare ti år siden var Fusarium-sopper et stort problem i norske kornavlinger. Noen viktige grep, og hell med været, har resultert i svært lave konsentrasjoner av mykotoksinet DON de siste årene.

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Aksfusariose er en kornsjukdom som kan angripe alle kornarter. Sjukdommen forårsakes av sopparter innen slekta Fusarium. Ulike Fusarium-arter kan produsere en rekke forskjellige mykotoksiner (soppgifter). Grenseverdier for innhold av enkelte mykotoksiner i korn og kornprodukter til mat og fôr er fastsatt av Mattilsynet (i henhold til EU’s regelverk). Denne dyrkningsveiledningen gir, på bakgrunn av dagens kunnskap, råd om hvordan en kan redusere risikoen for utvikling av mykotoksiner i korn.

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To mitigate the risk of erosion and nutrient runoff, reduced tillage has become more prevalent in Norway. Within within recent decades, there have been some years with relatively high occurrence of Fusarium head blight and mycotoxins in Norwegian cereal grain. This is thought to have been caused by an increased inoculum potential (IP) of Fusarium spp. due to larger amount of crop residues remaining on the soil surface, in combination with weather conditions promoting fungal growth and infection of cereal plants. The objective of this work was to elucidate the influence of different tillage practices on the IP of Fusarium spp. and the subsequent Fusarium-infection and mycotoxin contamination of spring wheat grain at harvest. Tillage trials were conducted at two locations in southeast Norway (Solør and Toten) over three years, 2010-2012. Residues of wheat from the previous year were collected in spring. Fusarium avenaceum and Fusarium graminearum were the most common Fusarium species recorded on wheat straw residues. IP was calculated as the percentage of the residues infested with Fusarium spp. multiplied by the proportion of the soil surface covered with residues. The IP of Fusarium spp. was lower in ploughed plots compared to those tilled with harrowing only. Ploughing in spring resulted in a similarly low IP as autumn ploughing. In contrast, harrowing in autumn generally reduced IP more than did spring harrowing. The mycotoxin levels in the harvested wheat were generally low, except for deoxynivalenol at high levels in Solør 2011. Despite a lower IP of ploughed versus harrowed plots, this was not reflected in the content of Fusarium and mycotoxins in harvested grain. The Fusarium species that dominated in the residues examined in this study were the same as those detected in the harvested grain, supporting the finding that residues are an important source of inoculum.

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The bread-making quality of wheat depends on the viscoelastic properties of the dough in which gluten proteins play an important role. The quality of gluten proteins is influenced by the genetics of the different wheat varieties and environmental factors. Occasionally, a near complete loss of gluten strength, measured as the maximum resistance towards stretching (Rmax), is observed in grain lots of Norwegian wheat. It is hypothesized that the loss of gluten quality is caused by degradation of gluten proteins by fungal proteases. To identify fungi associated with loss of gluten strength, samples from a selection of wheat grain lots with weak gluten (n = 10, Rmax < 0.3 N) and strong gluten (n = 10, Rmax ≥ 0.6 N) was analyzed for the abundance of fungal operational taxonomic units (OTUs) using DNA metabarcoding of the nuclear ribosomal Internal Transcribed Spacer (ITS) region ITS1. The DNA quantities for a selection of fungal pathogens of wheat, and the total amount of fungal DNA, were analyzed by quantitative PCR (qPCR). The mean level of total fungal DNA was higher in grain samples with weak gluten compared to grain samples with strong gluten. Heightened quantities of DNA from fungi within the Fusarium Head Blight (FHB) complex, i.e. Fusarium avenaceum, Fusarium graminearum, Microdochium majus, and Microdochium nivale, were observed in grain samples with weak gluten compared to those with strong gluten. Microdochium majus was the dominant fungus in the samples with weak gluten. Stepwise regression modeling based on different wheat quality parameters, qPCR data, and the 35 most common OTUs revealed a significant negative association between gluten strength and three OTUs, of which the OTU identified as M. majus was the most abundant. The same analysis also revealed a significant negative relationship between gluten strength and F. avenaceum detected by qPCR, although the DNA levels of this fungus were low compared to those of M. majus. In vitro growth rate studies of a selection of FHB species showed that all the tested isolates were able to grow with gluten as a sole nitrogen source. In addition, proteins secreted by these fungi in liquid cultures were able to hydrolyze gluten substrate proteins in zymograms, confirming their capacity to secrete gluten-degrading proteases. The identification of fungi with potential to influence gluten quality can enable the development of strategies to minimize future problems with gluten strength in food-grade wheat.

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Cereal grain contaminated by Fusarium mycotoxins is undesirable in food and feed because of the harmful health effects of the mycotoxins in humans and animals. Reduction of mycotoxin content in grain by cleaning and size sorting has mainly been studied in wheat. We investigated whether the removal of small kernels by size sorting could be a method to reduce the content of mycotoxins in oat grain. Samples from 24 Norwegian mycotoxin-contaminated grain lots (14 from 2015 and 10 from 2018) were sorted by a laboratory sieve (sieve size 2.2 mm) into large and small kernel fractions and, in addition to unsorted grain samples, analyzed with LC-MS-MS for quantification of 10 mycotoxins. By removing the small kernel fraction (on average 15% and 21% of the weight of the samples from the two years, respectively), the mean concentrations of HT-2+T-2 toxins were reduced by 56% (from 745 to 328 µg/kg) in the 2015 samples and by 32% (from 178 to 121 µg/kg) in the 2018 samples. Deoxynivalenol (DON) was reduced by 24% (from 191 to 145 µg/kg) in the 2018 samples, and enniatin B (EnnB) by 44% (from 1059 to 594 µg/kg) in the 2015 samples. Despite low levels, our analyses showed a trend towards reduced content of DON, ADON, NIV, EnnA, EnnA1, EnnB1 and BEA after removing the small kernel fraction in samples from 2015. For several of the mycotoxins, the concentrations were considerably higher in the small kernel fraction compared to unsorted grain. Our results demonstrate that the level of mycotoxins in unprocessed oat grain can be reduced by removing small kernels. We assume that our study is the first report on the effect of size sorting on the content of enniatins (Enns), NIV and BEA in oat grains.

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This study aims to understand the environmental factors, focusing on rain and fungal infection, affecting the assembly of glutenin polymers during grain maturation. Spring wheat was grown in the field and grains were sampled from 50% grain moisture until maturity. Grain moisture content, protein content, size of glutenin polymers, the presence of proteases, and the amount of DNA from common wheat pathogenic fungi were analysed. Rain influenced the rate of grain desiccation that occurred parallel to the rate of glutenin polymer assembly. Rapid desiccation contributed to faster glutenin polymer assembly than gradual desiccation. Severe reduction in the glutenin polymer size coincided with increased grain moisture due to rain. Furthermore, increased fungal DNA followed by presence of gluten-degrading proteases was observed in the grain after humid conditions. The presence of gluten-degrading proteases was presumably involved in reducing the size of glutenin polymers in grain. Our study gave new insight into how environmental conditions could be associated with the assembly of glutenin polymers during grain maturation. The results suggest that rain and/or fungal proteases play an important role in reducing the molecular size of glutenin polymers.

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Scientific communication is facilitated by a data-driven, scientifically sound taxonomy that considers the end-user's needs and established successful practice. Previously (Geiser et al. 2013; Phytopathology 103:400-408. 2013), the Fusarium community voiced near unanimous support for a concept of Fusarium that represented a clade comprising all agriculturally and clinically important Fusarium species, including the F. solani Species Complex (FSSC). Subsequently, this concept was challenged by one research group (Lombard et al. 2015 Studies in Mycology 80: 189-245) who proposed dividing Fusarium into seven genera, including the FSSC as the genus Neocosmospora, with subsequent justification based on claims that the Geiser et al. (2013) concept of Fusarium is polyphyletic (Sandoval-Denis et al. 2018; Persoonia 41:109-129). Here we test this claim, and provide a phylogeny based on exonic nucleotide sequences of 19 orthologous protein-coding genes that strongly support the monophyly of Fusarium including the FSSC. We reassert the practical and scientific argument in support of a Fusarium that includes the FSSC and several other basal lineages, consistent with the longstanding use of this name among plant pathologists, medical mycologists, quarantine officials, regulatory agencies, students and researchers with a stake in its taxonomy. In recognition of this monophyly, 40 species recently described as Neocosmospora were recombined in Fusarium, and nine others were renamed Fusarium. Here the global Fusarium community voices strong support for the inclusion of the FSSC in Fusarium, as it remains the best scientific, nomenclatural and practical taxonomic option available.

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Background Sphingolipids are structural components and signaling molecules in eukaryotic membranes, and many organisms produce compounds that inhibit sphingolipid metabolism. Some of the inhibitors are structurally similar to the sphingolipid biosynthetic intermediate sphinganine and are referred to as sphinganine-analog metabolites (SAMs). The mycotoxins fumonisins, which are frequent contaminants in maize, are one family of SAMs. Due to food and feed safety concerns, fumonisin biosynthesis has been investigated extensively, including characterization of the fumonisin biosynthetic gene cluster in the agriculturally important fungi Aspergillus and Fusarium. Production of several other SAMs has also been reported in fungi, but there is almost no information on their biosynthesis. There is also little information on how widely SAM production occurs in fungi or on the extent of structural variation of fungal SAMs. Results Using fumonisin biosynthesis as a model, we predicted that SAM biosynthetic gene clusters in fungi should include a polyketide synthase (PKS), an aminotransferase and a dehydrogenase gene. Surveys of genome sequences identified five putative clusters with this three-gene combination in 92 of 186 Fusarium species examined. Collectively, the putative SAM clusters were distributed widely but discontinuously among the species. We propose that the SAM5 cluster confers production of a previously reported Fusarium SAM, 2-amino-14,16-dimethyloctadecan-3-ol (AOD), based on the occurrence of AOD production only in species with the cluster and on deletion analysis of the SAM5 cluster PKS gene. We also identified SAM clusters in 24 species of other fungal genera, and propose that one of the clusters confers production of sphingofungin, a previously reported Aspergillus SAM. Conclusion Our results provide a genomics approach to identify novel SAM biosynthetic gene clusters in fungi, which should in turn contribute to identification of novel SAMs with applications in medicine and other fields. Information about novel SAMs could also provide insights into the role of SAMs in the ecology of fungi. Such insights have potential to contribute to strategies to reduce fumonisin contamination in crops and to control crop diseases caused by SAM-producing fungi.

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High concentrations of the mycotoxins HT-2 and T-2 (HT2 + T2), primarily produced by Fusarium langsethiae, have occasionally been detected in Norwegian oat grains. In this study, we identified weather variables influencing accumulation of HT2 + T2 in Norwegian oat grains. Oat grain samples from farmers’ fields were collected together with weather data (2004–2013). Spearman rank correlation coefficients were calculated between the HT2 + T2 contamination in oats at harvest and a range of weather summarisations within estimated phenological windows of growth stages in oats (tillering, flowering etc.). Furthermore, we developed a mathematical model to predict the risk of HT2 + T2 in oat grains. Our data show that adequate predictions of the risk of HT2 + T2 in oat grains at harvest can be achieved, based upon weather data observed during the growing season. Humid and cool conditions, in addition to moderate temperatures during booting, were associated with increased HT2 + T2 accumulation in harvested oat grains, whereas warm and humid weather during stem elongation and inflorescence emergence, or cool weather and absence of rain during booting reduced the risk of HT2 + T2 accumulation. Warm and humid weather immediately after flowering increased the risk, while moderate to warm temperatures and absence of rain during dough development, reduced the risk of HT2 + T2 accumulation in oat grains. Our data indicated that HT2 + T2 contamination in oats is influenced by weather conditions both pre- and post-flowering. These findings are in contrast with a previous study examining the risk of deoxynivalenol contamination in oat reporting that toxin accumulation was mostly influenced by weather conditions from flowering onwards.

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The natural occurrence of fungi, mycotoxins and fungal metabolites was investigated in 100 samples of maize grains collected from south and southwestern Ethiopia in 2015. The maize samples were contaminated by Fusarium, Aspergillus and Penicillium species. Using liquid chromatography tandem mass spectrometry 127 secondary metabolites were analysed. Zearalenone was the most prevalent mycotoxin, occurring in about 96% of the samples. Zearalenone sulfate was the second most prevalent, present in 81% of the samples. Fumonisin B1 was detected in 70% of the samples with a mean level of 606 μg kg−1 in positive samples, while FB2, FB3 and FB4 were detected in 62%, 51% and 60% of the maize samples with mean levels of 202, 136 and 85 μg kg−1, respectively. Up to 8% of the samples were contaminated with aflatoxins, with a maximum level of aflatoxin B1 of 513 μg kg−1. Results were higher than earlier reports for maize from Ethiopia.

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Production of chrysogine has been reported from several fungal genera including Penicillium, Aspergillus, and Fusarium. Anthranilic acid and pyruvic acid, which are expected precursors of chrysogine, enhance production of this compound. A possible route for the biosynthesis using these substrates is via a nonribosomal peptide synthetase (NRPS). Through comparative analysis of the NRPSs from genome-sequenced producers of chrysogine we identified a candidate NRPS cluster comprising five additional genes named chry2–6. Deletion of the two-module NRPS (NRPS14 = chry1) abolished chrysogine production in Fusarium graminearum, indicating that the gene cluster is responsible for chrysogine biosynthesis. Overexpression of NRPS14 enhanced chrysogine production, suggesting that the NRPS is the bottleneck in the biosynthetic pathway.

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In fungi, distribution of secondary metabolite (SM) gene clusters is often associated with host- or environment-specific benefits provided by SMs. In the plant pathogen Alternaria brassicicola (Dothideomycetes), the DEP cluster confers an ability to synthesize the SM depudecin, a histone deacetylase inhibitor that contributes weakly to virulence. The DEP cluster includes genes encoding enzymes, a transporter, and a transcription regulator. We investigated the distribution and evolution of the DEP cluster in 585 fungal genomes and found a wide but sporadic distribution among Dothideomycetes, Sordariomycetes, and Eurotiomycetes. We confirmed DEP gene expression and depudecin production in one fungus, Fusarium langsethiae. Phylogenetic analyses suggested 6–10 horizontal gene transfers (HGTs) of the cluster, including a transfer that led to the presence of closely related cluster homologs in Alternaria and Fusarium. The analyses also indicated that HGTs were frequently followed by loss/pseudogenization of one or more DEP genes. Independent cluster inactivation was inferred in at least four fungal classes. Analyses of transitions among functional, pseudogenized, and absent states of DEP genes among Fusarium species suggest enzyme-encoding genes are lost at higher rates than the transporter (DEP3) and regulatory (DEP6) genes. The phenotype of an experimentally-induced DEP3 mutant of Fusarium did not support the hypothesis that selective retention of DEP3 and DEP6 protects fungi from exogenous depudecin. Together, the results suggest that HGT and gene loss have contributed significantly to DEP cluster distribution, and that some DEP genes provide a greater fitness benefit possibly due to a differential tendency to form network connections.

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Putative proton coupled di-peptide transporters, PTR2s, are found in filamentous fungi in different numbers and their function during fungal development and plant infection is unresolved. In Fusarium graminearum, the cause of head blight in cereals, we identified four putative PTR2 transporters (FgPTR2A-D). The genes did not cluster together in phylogenetic analyses and only FgPTR2A and FgPTR2C were able to complement a PTR2 deficient yeast mutant in uptake of di-peptides. All FgPTR2s are continuously expressed throughout the fungal lifecycle, although at different levels. In silico analyses of existing expression-data show that FgPTR2B is found at higher levels than the others in planta and during sexual development. Deletion mutants of FgPTR2A, FgPTR2C, and FgPTR2D had a higher production of deoxynivalenol (DON) and zearalenone and lower production of fusarielin H than the wild type. Perithecium development was reduced in these mutants but unaffected by deletion of FgPTR2B. Conidia production was reduced in the FgPTR2B mutant and unaffected by deletion of the other PTR2 transporters. Sexual development and secondary metabolite production are known to be linked at the regulatory level and the results suggest that PTR2s are active in nitrogen turnover and thereby influence signal processes.

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Four field trials (spring wheat and oats) were conducted (one on clay soil, one on loam soil and two on silt soil) for three years in important cereal growing districts, to investigate the influence of tillage regimes (ploughing versus reduced tillage in either autumn or spring) and straw management (removed and retained) on plant residue amounts, weed populations, soil structural parameters and cereal yields. The effect of tillage on soil structure varied, mainly due to the short trial period. In general, the amount of small soil aggregates increased with tillage intensity. Reduced soil tillage, and in some cases spring ploughing, gave significantly higher aggregate stability than autumn ploughing, thus providing protection against erosion. However, decreasing tillage intensity increased the amounts of weeds, particularly of Poa annua on silt soil. Straw treatment only slightly affected yields, while effects of tillage varied between both year and location. Reduced tillage, compared to ploughing, gave only small yield differences on loam soil, while it was superior on clay soil and inferior on silt soil. Our results suggest that shallow spring ploughing is a good alternative to autumn ploughing, since it gave comparable yields, better protection against erosion and was nearly as effective against weeds.

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The mycotoxin enniatin B, a cyclic hexadepsipeptide produced by the plant pathogen Fusarium, is prevalent in grains and grain-based products in different geographical areas. Although enniatins have not been associated with toxic outbreaks, they have caused toxicity in vitro in several cell lines. In this study, the cytotoxic effects of enniatin B were assessed in relation to cellular energy metabolism, cell proliferation, and the induction of apoptosis in Balb 3T3 and HepG2 cells. The mechanism of toxicity was examined by means of whole genome expression profiling of exposed rat primary hepatocytes. Enniatin B altered cellular energy metabolism and reduced cell proliferation in Balb 3T3 and HepG2 cell lines. Furthermore, the proportion of apoptotic cell populations of Balb 3T3 cells slightly increased. On the other hand, enniatin B caused necrotic cell death in primary hepatocytes. Gene expression studies revealed the alteration of energy metabolism due to effects on mitochondrial organization and function and the assembly of complex I of the electron transport chain.

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High concentrations of the mycotoxin deoxynivalenol (DON), produced by Fusarium graminearum have occurred frequently in Norwegian oats recently. Early prediction of DON levels is important for farmers, authorities and the Cereal Industry. In this study, the main weather factors influencing mycotoxin accumulation were identified and two models to predict the risk of DON in oat grains in Norway were developed: (1) as a warning system for farmers to decide if and when to treat with fungicide, and (2) for authorities and industry to use at harvest to identify potential food safety problems. Oat grain samples from farmers’ fields were collected together with weather data (2004–2013). A mathematical model was developed and used to estimate phenology windows of growth stages in oats (tillering, flowering etc.). Weather summarisations were then calculated within these windows, and the Spearman rank correlation factor calculated between DON-contamination in oats at harvest and the weather summarisations for each phenological window. DON contamination was most clearly associated with the weather conditions around flowering and close to harvest. Warm, rainy and humid weather during and around flowering increased the risk of DON accumulation in oats, as did dry periods during germination/seedling growth and tillering. Prior to harvest, warm and humid weather conditions followed by cool and dry conditions were associated with a decreased risk of DON accumulation. A prediction model, including only pre-flowering weather conditions, adequately forecasted risk of DON contamination in oat, and can aid in decisions about fungicide treatments.

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A glutathione (GSH) adduct of the mycotoxin 4-deoxynivalenol (DON), together with a range of related conjugates, has recently been tentatively identified by LC-MS of DON-treated wheat spikelets. In this study, we prepared samples of DON conjugated at the 10- and 13-positions with GSH, Cys, CysGly, -GluCys and N-acetylcysteine (NAC). The mixtures of conjugates were used as standards for LC-HRMS analysis of one of the DON-treated wheat spikelet samples, as well as 19 Norwegian grain samples of spring wheat and 16 grain samples of oats that were naturally-contaminated with DON at concentrations higher than 1 mg/kg. The artificially-contaminated wheat spikelets contained conjugates of GSH, CysGly and Cys coupled at the olefinic 10-position of DON, whereas the naturally-contaminated harvest-ripe grain samples contained GSH, CysGly, Cys, and NAC coupled mainly at the 13-position on the epoxy group. The identities of the conjugates were confirmed by LC-HRMS comparison with authentic standards, oxidation to the sulfoxides with hydrogen peroxide, and examination of product-ion spectra from LC-HRMS/MS analysis. No -GluCys adducts of DON were detected in any of the samples. The presence of 15-O-acetyl-DON was demonstrated for the first time in Norwegian grain. The results indicate that a small but significant proportion of DON is metabolized via the GSH-conjugation pathway in plants. To our knowledge, this is the first report of in vivo conjugation of trichothecenes via their epoxy group, which has generally been viewed as unreactive. Because conjugation at the 13-position of DON and other trichothecenes has been shown to be irreversible, this type of conjugate may prove useful as a biomarker of exposure to DON and other 12,13-epoxytrichothecenes.

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This paper presents peer-reviewed studies comparing the content of deoxynivalenol (DON), HT-2+T-2 toxins, zearalenone (ZEA), nivalenol (NIV), ochratoxin A (OTA) and fumonisins in cereal grains, and patulin (PAT) in apple and apple-based products, produced in organically and conventionally grown crops in temperate regions. Some of the studies are based on data from controlled field trials, however, most are farm surveys and some are food basket surveys. Almost half of the studies focused on DON in cereals. The majority of these studies found no significant difference in DON content in grain from the two farming systems, but several studies showed lower DON content in organically than in conventionally produced cereals. A number of the investigations reported low DON levels in grain, far below the EU limits for food. Many authors suggested that weather conditions, years, locations, tillage practice and crop rotation are more important for the development of DON than the type of farming. Organically produced oats contained mainly lower levels of HT-2+T-2 toxins than conventionally produced oats. Most studies on ZEA reported no differences between farming systems, or lower concentrations in organically produced grain. For the other mycotoxins in cereals, mainly low levels and no differences between the two farming systems were reported. Some studies showed higher PAT contamination in organically than in conventionally produced apple and apple products. The difference may be due to more efficient disease control in conventional orchards. It cannot be concluded that any of the two farming systems increases the risk of mycotoxin contamination. Despite no use of fungicides, an organic system appears generally able to maintain mycotoxin contamination at low levels. More systematic comparisons from scientifically controlled field trials and surveys are needed to clarify if there are differences in the risk of mycotoxin contamination between organically and conventionally produced crops.

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Lack of resistance to pink snow mould (Microdochium nivale) is a major constraint for adaptation of perennial ryegrass (Lolium perenne L.) to continental regions with long-lasting snow cover at higher latitudes. Almost all investigations of genetic variation in resistance have been performed using cold acclimated plants. However, there may be variation in resistance mechanisms that are functioning independently of cold acclimation. In this study our aim was to identify candidate genes involved in such resistance mechanisms. We first characterized variation in resistance to M. nivale among non-acclimated genotypes from the Norwegian cultivar ‘Fagerlin’ based on relative regrowth and fungal quantification by real-time qPCR. One resistant and one susceptible genotype were selected for transcriptome analysis using paired-end sequencing by Illumina Hiseq 2000. Transcriptome profiles, GO enrichment and KEGG pathway analysis indicate that defense response related genes are differentially expressed between the resistant and the susceptible genotype. A significant up-regulation of defense related genes, as well as genes involved in cell wall cellulose metabolic processes and aryl-alcohol dehydrogenase (NADP+) activity, was observed in the resistant genotype. The candidate genes identified in this study might be potential molecular marker resources for breeding perennial ryegrass cultivars with improved resistance to pink snow mould.

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Biosynthesis of the black perithecial pigment in the filamentous fungus Fusarium graminearum is dependent on the polyketide synthase PGL1 (oPKS3). A seven-membered PGL1 gene cluster was identified by over-expression of the cluster specific transcription factor pglR. Targeted gene replacement showed that PGL1, pglJ, pglM and pglV were essential for the production of the perithecial pigment. Over-expression of PGL1 resulted in the production of 6-O-demethyl-5-deoxybostrycoidin (1), 5-deoxybostrycoidin (2), and three novel compounds 5-deoxybostrycoidin anthrone (3), 6-O-demethyl-5-deoxybostrycoidin anthrone (4) and purpurfusarin (5). The novel dimeric bostrycoidin purpurfusarin (5) was found to inhibit the growth of Candida albicans with an IC50 of 8.0 +/− 1.9 μM. The results show that Fusarium species with black perithecia have a previously undescribed form of 5-deoxybostrycoidin based melanin in their fruiting bodies.

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During the last ten years, Norwegian cereal grain industry has experienced large challenges due to Fusarium spp. and Fusarium mycotoxin contamination of small-grained cereals. To prevent severely contaminated grain lots from entering the grain supply chain, it is important to establish surveys for the most prevalent Fusarium spp. and mycotoxins. The objective of our study was to quantify and calculate the associations between Fusarium spp. and mycotoxins prevalent in oats and spring wheat. In a 6-year period from 2004-2009, 178 grain samples of spring wheat and 289 samples of oats were collected from farmers’ fields in South East Norway. The grains were analysed for 18 different Fusarium-mycotoxins by liquid chromatography – mass spectrometry. Generally, the median mycotoxin levels were higher than reported in Norwegian studies covering previous years. The DNA content of Fusarium graminearum, Fusarium culmorum, Fusarium langsethiae, Fusarium poae and Fusarium avenaceum were determined by quantitative PCR. We identified F. graminearum as the main deoxynivalenol (DON) producer in oats and spring wheat, and F. langsethiae as the main HT-2 and T-2-toxins producer in oats. No association was observed between quantity of F. graminearum DNA and quantity of F. langsethiae DNA nor for their respective mycotoxins, in oats. F. avenaceum was one of the most prevalent Fusarium species in both oats and spring wheat. The following ranking of Fusarium species was made based on the DNA concentrations of the Fusarium spp. analysed in this survey (from high to low): F. graminearum = F. langsethiae = F. avenaceum > F. poae > F. culmorum (oats); F. graminearum = F. avenaceum > F. culmorum > F. poae = F. langsethiae (spring wheat). Our results are in agreement with recently published data indicating a shift in the relative prevalence of Fusarium species towards more F. graminearum versus F. culmorum in Norwegian oats and spring wheat.

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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.

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Hvilken jordarbeiding som benyttes i den enkelte kornåker påvirker blant annet avlingsmengde, kvaliteten på kornet og miljøet. I denne publikasjonen er det samlet informasjon om effekter av ulik jordarbeiding, som hjelp til korndyrkere ved vurdering av jordarbeidingsmetoder, og for myndigheter ved beslutninger om jordarbeiding i regionale miljøprogram (RMP).

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In this letter, we advocate recognizing the genus Fusarium as the sole name for a group that includes virtually all Fusarium species of importance in plant pathology, mycotoxicology, medicine, and basic research. This phylogenetically guided circumscription will free scientists from any obligation to use other genus names, including teleomorphs, for species nested within this clade, and preserve the application of the name Fusarium in the way it has been used for almost a century. Due to recent changes in the International Code of Nomenclature for algae, fungi, and plants, this is an urgent matter that requires community attention. The alternative is to break the longstanding concept of Fusarium into nine or more genera, and remove important taxa such as those in the F. solani species complex from the genus, a move we believe is unnecessary. Here we present taxonomic and nomenclatural proposals that will preserve established research connections and facilitate communication within and between research communities, and at the same time support strong scientific principles and good taxonomic practice.

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Plogen har tradisjonelt vært viktig for å få et godt såbed, for innblanding av halmrester og gjødsel i jorda, og for god bekjemping av ugras og sjukdommer. Redusert jordarbeiding uten bruk av plogen, gir imidlertid store miljøfordeler i form av mindre erosjon og utvasking av næringsstoffer. Denne rapporten fokuserer på konsekvenser av ulik jordarbeiding på plantevernsituasjonen i korn. Basert på dagens kunnskap fra norske og internasjonale studier konkluderes det med at redusert jordarbeiding gir økt risiko for utvikling av ugras og plantesjukdommer, samt mykotoksiner. I tillegg kan redusert jordarbeiding føre til økt bruk av kjemiske plantevernmidler som glyfosat, fenoksysyrer og soppmidler. Ugraset og de fleste plantesjukdommer kan som regel bekjempes med plantevernmidler, mens Fusarium spp. og mykotoksiner bare delvis kan bekjempes av kjemiske midler. Miljørisikoen av kjemiske plantevernmidler påvirkes av egenskaper til plantevernmidlene. Redusert jordarbeiding fører til økt risiko for transport til grunnvann av fenoksysyrer og lavdosemidler (sulfonylurea-preparater). Risiko for transport til overflatevann av ugrasmidler og soppmidler er minst når åkeren ligger i stubb. Været og klimaet har stor betydning for utvikling av skadegjørerne, risiko for utvikling av mykotoksiner og utvasking av plantevernmidler. Vårpløying kan være gunstig miljømessig sett fordi det kan redusere erosjon og næringsstofftap. Samtidig gir det mindre behov for, og derfor redusert miljørisiko av, plantevernmidler enn andre typer jordarbeiding. Vårpløying egner seg derimot dårlig på stiv leirjord, det fører til større tidspress på våren og risiko for forsinka våronn og derved lavere avling.

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Aksfusariose er en utbredt og destruktiv sjukdom i korn som kan forårsakes av en rekke ulike sopparter innen slekta Fusarium. I tillegg til å redusere avlingsmengde, kvalitet og frøspiring, kan ulike Fusarium-arter produsere en rekke ulike soppgifter (mykotoksiner) som kan være giftige for mennesker og dyr. Fuktige værforhold i perioden rundt blomstring av kornet ser ut til å øke risikoen for angrep av Fusarium. I tillegg kan dyrkningspraksis påvirke forekomsten av aksfusariose og utvikling av mykotoksiner i kornet.

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Fusarium langsethiae is a recently characterized fungus within the genus Fusarium. It is found as a grain contaminant of small grain cereals such as oats and barley, and to a lesser extent wheat. Fusarium langsethiae is particularly widespread in the Nordic countries and the UK where it poses a serious problem as the main producer of T-2 and HT-2 mycotoxins. The biology of F. langsethiae and its interaction with the plant remains poorly understood, partly hampered by difficulties reproducing a natural level of infection under controlled conditions. The reported study was designed as a series of glasshouse experiments to advance our understanding of F. langsethiae biology by investigating alternative infection routes and its proliferation in oats, Avena sativa. Various methods of seed, soil, and seedling inoculation, boot injection and spray inoculation, were tested. The results clearly show a strong preference of F. langsethiae for the panicle, ruling out alternative infection routes. At relatively low temperatures spray infection, accompanied by prolonged humidity, ensured a thorough establishment of the fungus both at flowering and at early dough stage. Boot injection proved to be a reliable working tool for production of an even and predictable grain infection. Apart from in the panicle, considerable fungal proliferation was only detected in flag leaf nodes, and was a direct consequence of the boot injection method. Fungal presence in the node tissue also correlated with significant stunting of infected shoots. In light of the results the pathogenic and endophytic abilities of F. langsethiae are discussed.

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Gårdsvarsling gjør det mulig å beregne lokalt tilpassede plantevernvarsler ved bruk av værvarsel og radarmåling av nedbør. Vanligvis beregnes plantevernvarsler på grunnlag av data fra værstasjoner, men mange gårdsbruk ligger langt unna nærmeste målepunkt. For disse brukene vil varsler basert på værvarsler og radarmålt nedbør være et godt alternativ. Gårdsvarsler tilpasset det enkelte gårdsbruk vil derfor øke nytteverdien av varslingstjenesten innen planteskadegjørere (VIPS) for mange dyrkere.

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Aksfusariose er en kornsjukdom som kan angripe alle kornarter. Sjukdommen forårsakes av sopparter innen slekta Fusarium. Ulike Fusarium-arter kan produsere en rekke forskjellige mykotoksiner (soppgifter). Grenseverdier for innhold av enkelte mykotoksiner i korn og kornprodukter til mat og fôr er fastsatt av Mattilsynet (i henhold til EU’s regelverk). Denne dyrkningsveiledningen gir, på bakgrunn av dagens kunnskap, råd om hvordan en kan redusere risikoen for utvikling av mykotoksiner i korn.

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The effect of potential resistance inducing chemicals on disease development of Fusarium head blight was studied in winter wheat (Triticum aestivum L.). As a pre-screening test, the effect of different treatments on development of Microdochium majus (syn. Microdochium nivale var. majus) was studied in detached leaves. Based on these tests, DL-3-aminobutyric acid, Bion (benzo-(1,2,3) thiadiazole-7-carbothioic acid S-methyl ester), and a foliar fertilizer containing potassium phosphite were selected for further studies. Greenhouse-grown winter wheat was sprayed with aqueous solutions of the potential resistance inducers 7 days prior to Fusarium culmorum point inoculation of the heads. Disease development was registered as number of bleached spikelets per inoculated spike. Spraying plants with the foliar fertilizer reduced the disease severity of F. culmorum by up to 40%. A reduced disease development of M. majus was also observed in detached leaves pre-treated with the foliar fertilizer. When the foliar fertilizer was added to the growth medium, a reduced in vitro growth of M. majus and F. culmorum was observed, indicating that the effect on disease development is at least partly due to a fungistatic effect. No significant reduction in disease development was observed in wheat pre-treated with DL-3-aminobutyric acid or Bion, although these compounds tended to reduce disease development, especially when applied in combination with other potential resistance inducers. We conclude that spraying winter wheat with a solution containing potassium phosphite can reduce development of M. majus and F. culmorum.

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The estrogenic mycotoxin zearalenone (ZON) produced by some Fusarium spp. causes reproductive problems and hyperestrogenic syndromes in mammals. In an effort to elucidate the molecular pathways of ZON production, we present a comparative real-time quantitative polymerase chain reaction expression study of seven contiguous genes in the ZON biosynthetic cluster on sterile rice and during wheat and oat infection. Under ZON production on rice, the polyketide synthase (PKS) genes PKS4 and PKS13, alcohol oxidase FG12056 gene, and transcriptional regulator FG02398 gene showed similarly upregulated patterns, whereas the nonribosomal peptide synthetase (NPS) FG02394, the K+ channel beta subunit FG12015, and the protein kinase FG02399 displayed a variant pattern. During the same time period under wheat infection when no ZON was produced, the PKS genes and the NPS were downregulated relative to rice whereas the K+ channel beta subunit gene FG12015 was markedly upregulated, suggesting that it may play a role in the infection process. This is the first expression study of ZON biosynthetic genes in planta. The results give insight into the regulation and activities of the ZON gene cluster under different experimental systems and suggest a connection between ZON and a K+ channel that could reveal a novel function for ZON in Fusarium spp.

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The Fusarium genus includes devastating plant pathogenic fungi that cause diseases in cereals around the world. They produce several mycotoxins, including the estrogenic compound zearalenone. To better understand the molecular mechanisms determining zearalenone production, we performed differential display RT-PCR under conditions where Fusarium graminearum and F. culmorum produced high amounts of zearalenone. We found 133 expressed sequence tags (ESTs) and 54 of these were considered to be up-regulated during high zearalenone production. Several of the ESTs were confirmed to be up-regulated by real-time qPCR, but none showed any significant down-regulation in the zearalenone negative mutant Delta PKS4-T9, or were similar to typical gene expression patterns of previously described zearalenone-related genes. Some of the up-regulated ESTs were similar to genes involved in secondary metabolite production, lipid metabolism, transcriptional activation, provision of precursors, signal transduction, transport or detoxification. Several of the ESTs were also located adjacent to one another in the genome and therefore might represent genes involved in the same biosynthetic pathway. Members of six such putative pathways could be found. All sequences were compared to the MIPS F. graminearum Genome Database to verify autocalled gene predictions experimentally and to introduce new exons and gene structures.

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Zearalenones are produced by several Fusarium species and can cause reproductive problems in animals. Some aurofusarin mutants of Fusarium pseudograminearum produce elevated levels of zearalenone (ZON), one of the estrogenic mycotoxins comprising the zearalenones. An analysis of transcripts from polyketide synthase genes identified in the Fusarium graminearum database was carried out for these mutants. PKS4 was the only gene with an enoyl reductase domain that had a higher level of transcription in the aurofusarin mutants than in the wild type. An Agrobacterium tumefaciens-mediated transformation protocol was used to replace the central part of the PKS4 gene with a hygB resistance gene through double homologous recombination in an F. graminearum strain producing a high level of ZON. PCR and Southern analysis of transformants were used to identify isolates with single insertional replacements of PKS4. High-performance liquid chromatography analysis showed that the PKS4 replacement mutant did not produce ZON. Thus, PKS4 encodes an enzyme required for the production of ZON in F. graminearum. Barley root infection studies revealed no alteration in the pathogenicity of the PKS4 mutant compared to the pathogenicity of the wild type. The expression of PKS13, which is located in the same cluster as PKS4, decreased dramatically in the mutant, while transcription of PKS4 was unchanged. This differential expression may indicate that ZON or its derivatives do not regulate expression of PKS4 and that the PKS4-encoded protein or its product stimulates expression of PKS13. Furthermore, both the lack of aurofusarin and ZON influenced the expression of other polyketide synthases, demonstrating that one polyketide can influence the expression of others.

Prosjekter

20030730ef0198-korn-utdrag
Adaptations within the Norwegian wheat value chain to improve quality and obtain high and stable quantities for milling in the forthcoming decades (MATHVETE)


This project aims to improve the quality of Norwegian wheat used for milling to secure high and stable production in forthcoming decades under more challenging climatic conditions. Increasing wheat production for milling is the most efficient way to achieve increased domestic food production in Norway and it will strengthen the competitiveness in the agricultural sector.

Active Updated: 10.12.2020
End: mars 2023
Start: jan 2019
bilde1

Divisjon for bioteknologi og plantehelse

PlantStrength – Strenghtening the basis of sound plant protection

This strategic institute initiative aims to understand the interactions between plant pathogens, invertebrate pests, weeds and natural enemies to evaluate and improve management strategies for cereals under the current and future Norwegian climate. The work is divided in 3 work packages: WP1: The Fusarium-Oat system WP2: The Cereal pest syndrome WP 3: Simulation computer models.

Finished Updated: 23.09.2017
End: des 2016
Start: jan 2012