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the trichothecene biosynthetic pathway.

Molecular and genetic studies of Fusarium trichothecene biosynthesis: pathways, genes, and evolution.

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Citrinin is a mycotoxin which is often found in food

ERGOT III.1 Properties and analytical methods III.1.1 Chemical properties III.1.2 Analytical methods for ergot and ergot alkaloids III.1.2.1 Ergot III.1.2.2 Ergot alkaloids III.2 Sources and occurrence III.2.1 Fungal producers III.2.2 Biosynthesis III.2.3 Occurrence in foodstuffs III.2.4 Fate of ergolines during food processing III.3 Metabolism III.4 Effects on animals III.4.1 Field studies III.4.2 Experimental animal studies III.4.2.1 Cattle III.4.2.2 Sheep III.4.2.3 Poultry III.4.2.4 Swine III.4.2.5 Primates III.5 Effects on man III.5.1 Ergometrine-related outbreaks III.5.2 Clavine-related outbreaks III.6 Evaluation of the human health risks REFERENCESRESUMERESUMENWHO TASK GROUP ON SELECTED MYCOTOXINS: OCHRATOXINS, TRICHOTHECENES, AND ERGOT MembersProfessor W.W.

Our results show that trichothecene biosynthesis involves a complex developmental process that ..

Ancymidol, a plant growth regulator, inhibited biosynthesis of diacetoxyscirpenol by Gibberella pulicaris (Fusarium sambucinum) in a defined liquid medium. Ancymidol also inhibited biosynthesis of T-2 toxin by a wild-type strain of Fusarium sporotrichioides and biosynthesis of diacetoxyscirpenol, deacetylated calonectrin, and dideacetylated calonectrin by mutant strains of this species. Ancymidol-treated cultures accumulated the hydrocarbon trichodiene, a biosynthetic precursor of the trichothecenes. Ancymidol did not block trichodiene accumulation by a trichodiene-producing mutant strain of F. sporotrichioides. Ancymidol appears to block the trichothecene biosynthetic pathway after formation of trichodiene and before formation of trichothecenes containing four or more oxygen atoms.

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Tri genes expression was determined by using real time-PCR and RT-PCR and confirmed the expression of Tri5, Tri6 and Tri12 among three genotypes. The comparison of genotypes and time points showed that, in Gamenya, the expression of Tri5 was increased significantly at 7 dai (t-test, pF. g virulence during the infection stage depends on thrichothecene or mycotoxin release from the pathogen, for that reason we determined the expression of Tri12 by using RT-PCR (Figure 5). In Gamenya and Sumai 3 at both time points found their expression, whereas in Nobeokabouzu-komugi at 7 dai there was no expression. This result demonstrated trichothecene biosynthesis and the product (mycotoxin) released in Gamenya and Sumai 3, but in Nobeokabouzu the trichothecene biosynthesis were reduced and the product released were not active at 7 dai. One possibility is that the expressions of defense-related genes in this highly resistant cultivar are active that inhibit the trichothecene release and provoked toxicity to the fungus itself. However, in order to confirm this speculation it is important to quantify trichothecene (mycotoxin) content in these cultivars. In our laboratory previously determined the DON content in Gamenya, Sumai 3 and Nobeokabouzu at 10 dai and the result clearly showed that DON content was high in Gamenya and less in Sumai 3 and Nobeokabouzu being 38.155, 13.500 and 10.955 ppm, respectively.

TRI11 is the fifth gene to be identified within the trichothecene pathway gene cluster of F. sporotrichioides. On the basis of a restriction map we constructed for cosmid 9-1 (Fig. ), TRI11 is located approximately 3.8 kb from TRI5 and is transcribed in the opposite direction. Disruption of TRI11 results in an altered trichothecene profile that is consistent with the participation of the TRI11 gene product in trichothecene biosynthesis. Transformants lacking a functional copy of TRI11 do not accumulate T-2 toxin and other late-pathway trichothecenes characteristic of F. sporotrichioides NRRL 3299 but instead accumulate the earlier-pathway intermediate isotrichodermin. Finally, expression of TRI11 is coincident with the expression of other pathway genes, as shown by the detection of a TRI11 transcript in RNA isolated from cultures grown under conditions known to support the expression of other pathway genes ().

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In FHB susceptible cultivar Gamenya, fungal biomass increased at 7 dai, whereas in resistant cultivar Sumai 3 it was reduced (Figure 1). We expected minimum F. g biomass in highly resistant cultivar Nobeokabouzu-komugi, however, it increased drastically at 7 dai. We assume that in susceptible cv. Gamenya, fungus biomass is less due to the less availability of nutrients required for the pathogen development when compare to resistant cultivars. Another reason might be the less requirement of fungus activity to infect susceptible cultivar necrotized tissue (Figure 2), however, in resistant cultivars (symptomless) required abundant fungus in order to suppress host defense genes, colonize and develop the disease. Here hypothesized that the F. g biomass does not have direct relation with fungal disease symptom, and depends on pathogen virulence as well as expression of virulence genes. In order to clarify the relation between fungus virulence gene expression and disease symptoms, expression of trichothecene biosynthesis genes (Tri genes) such as Tri5, Tri6 and Tri12 in both time points was determined.

To isolate RNA, cultures were grown in GYEP medium for 23 h and harvested by filtration. These growth conditions support trichothecene biosynthesis and result in high-level expression of several pathway gene mRNAs (). The mycelial mats (approximately 0.5 to 1.0 g) were immediately ground in liquid N2, and RNA was isolated with an RNaid kit (Bio 101) by the acid-phenol procedure described in the manufacturer’s product literature. Northern blotting was carried out as described elsewhere () by using a 32P-labeled probe made from the SacI-HindIII fragment cloned in pFSC3-6.

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  • Ancymidol blocks trichothecene biosynthesis and leads …

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  • Oxygenation steps in trichothecene biosynthesis are of ..

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Nutrient profiling reveals potent inducers of ..

The trichothecene biosynthetic pathway begins with the cyclization of farnesyl pyrophosphate by the enzyme trichodiene synthase to form trichodiene (Fig. ). Trichodiene undergoes multiple oxygenations involving molecular oxygen and as many as four different esterifications () to form trichothecene products, such as T-2 toxin. Like other fungal terpenoids, the trichothecenes are produced as a large family of structurally related compounds, with individual Fusarium species typically producing distinctive trichothecene profiles. Oxygenation steps in trichothecene biosynthesis are of particular interest since the degree of oxygenation greatly alters trichothecene toxicity. At least one trichothecene oxygen, the 12,13-epoxide, is required for toxicity (). In Fusarium sporotrichioides, the initial oxygenation of trichodiene in the biosynthesis of T-2 toxin (Fig. ) is catalyzed by a cytochrome P-450 monooxygenase encoded by TRI4 (). Hydroxylation of C-15 is the first oxygenation step employing an intermediate containing the core trichothecene structure (). The product of the C-15 hydroxylase, 15-decalonectrin, is most likely the last common intermediate of the various Fusarium trichothecene pathways. Biosynthesis of deoxynivalenol, the trichothecene most commonly detected in grains, is thought to branch off of the T-2 toxin pathway at this intermediate.

Comparison of Trichothecene Biosynthetic Gene …

Several genes in the trichothecene biosynthetic pathway of Fusarium sporotrichioides have been shown to reside in a gene cluster. Sequence analysis of a cloned DNA fragment located 3.8 kb downstream from TRI5 has led to the identification of the TRI11 gene. The nucleotide sequence of TRI11 predicts a polypeptide of 492 residues (Mr = 55,579) with significant similarity to members of the cytochrome P-450 superfamily. TRI11 is most similar to several fungal cytochromes P-450 (23 to 27% identity) but is sufficiently distinct to define a new cytochrome P-450 gene family, designated CYP65A1. Disruption of TRI11 results in an altered trichothecene production phenotype characterized by the accumulation of isotrichodermin, a trichothecene pathway intermediate. The evidence suggests that TRI11 encodes a C-15 hydroxylase involved in trichothecene biosynthesis.

PLOS ONE: Cellular Development Associated with …

From this work, we propose that products of the arginine-polyamine biosynthetic pathway in plants may play a role in the induction of trichothecene biosynthesis during infection.

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