Fusarium sporotrichioides Produces Two HT-2-α-Glucosides on Rice.

Fusarium is a genus that mostly consists of plant pathogenic fungi which are able to produce a broad range of toxic secondary metabolites. In this study, we focus on a type A trichothecene-producing isolate (15-39) of Fusarium sporotrichioides from Lower Austria. We assessed the secondary metabolite profile and optimized the toxin production conditions on autoclaved rice and found that in addition to large amounts of T-2 and HT-2 toxins, this strain was able to produce HT-2-glucoside. The optimal conditions for the production of T-2 toxin, HT-2 toxin, and HT-2-glucoside on autoclaved rice were incubation at 12 °C under constant light for four weeks, darkness at 30 °C for two weeks, and constant light for three weeks at 20 °C, respectively. The HT-2-glucoside was purified, and the structure elucidation by NMR revealed a mixture of two alpha-glucosides, presumably HT-2-3-O-alpha-glucoside and HT-2-4-O-alpha-glucoside. The efforts to separate the two compounds by HPLC were unsuccessful. No hydrolysis was observed with two the alpha-glucosidases or with human salivary amylase and Saccharomyces cerevisiae maltase. We propose that the two HT-2-alpha-glucosides are not formed by a glucosyltransferase as they are in plants, but by a trans-glycosylating alpha-glucosidase expressed by the fungus on the starch-containing rice medium.

The Role of Plant Hormones in the Interaction of Colletotrichum Species with Their Host Plants.

Colletotrichum is a plant pathogenic fungus which is able to infect virtually every economically important plant species. Up to now no common infection mechanism has been identified comparing different plant and Colletotrichum species. Plant hormones play a crucial role in plant-pathogen interactions regardless whether they are symbiotic or pathogenic. In this review we analyze the role of ethylene, abscisic acid, jasmonic acid, auxin and salicylic acid during Colletotrichum infections. Different Colletotrichum strains are capable of auxin production and this might contribute to virulence. In this review the role of different plant hormones in plant-Colletotrichum interactions will be discussed and thereby auxin biosynthetic pathways in Colletotrichum spp. will be proposed.

Identification and Functional Characterization of the Gene Cluster Responsible for Fusaproliferin Biosynthesis in Fusarium proliferatum.

The emerging mycotoxin fusaproliferin is produced by Fusarium proliferatum and other related Fusarium species. Several fungi from other taxonomic groups were also reported to produce fusaproliferin or the deacetylated derivative, known as siccanol or terpestacin. Here, we describe the identification and functional characterization of the Fusarium proliferatum genes encoding the fusaproliferin biosynthetic enzymes: a terpenoid synthase, two cytochrome P450s, a FAD-oxidase and an acetyltransferase. With the exception of one gene encoding a CYP450 (FUP2, FPRN_05484), knock-out mutants of the candidate genes could be generated, and the production of fusaproliferin and intermediates was tested by LC-MS/MS. Inactivation of the FUP1 (FPRN_05485) terpenoid synthase gene led to complete loss of fusaproliferin production. Disruption of a putative FAD-oxidase (FUP4, FPRN_05486) did not only affect oxidation of preterpestacin III to terpestacin, but also of new side products (11-oxo-preterpstacin and terpestacin aldehyde). In the knock-out strains lacking the predicted acetyltransferase (FUP5, FPRN_05487) fusaproliferin was no longer formed, but terpestacin was found at elevated levels. A model for the biosynthesis of fusaproliferin and of novel derivatives found in mutants is presented.

Biochemical Characterization of the Fusarium graminearum Candidate ACC-Deaminases and Virulence Testing of Knockout Mutant Strains.

Fusarium graminearum is a plant pathogenic fungus which is able to infect wheat and other economically important cereal crop species. The role of ethylene in the interaction with host plants is unclear and controversial. We have analyzed the inventory of genes with a putative function in ethylene production or degradation of the ethylene precursor 1-aminocyclopropane carboxylic acid (ACC). F. graminearum, in contrast to other species, does not contain a candidate gene encoding ethylene-forming enzyme. Three genes with similarity to ACC synthases exist; heterologous expression of these did not reveal enzymatic activity. The F. graminearum genome contains in addition two ACC deaminase candidate genes. We have expressed both genes in E. coli and characterized the enzymatic properties of the affinity-purified products. One of the proteins had indeed ACC deaminase activity, with kinetic properties similar to ethylene-stress reducing enzymes of plant growth promoting bacteria. The other candidate was inactive with ACC but turned out to be a d-cysteine desulfhydrase. Since it had been reported that ethylene insensitivity in transgenic wheat increased Fusarium resistance and reduced the content of the mycotoxin deoxynivalenol (DON) in infected wheat, we generated single and double knockout mutants of both genes in the F. graminearum strain PH-1. No statistically significant effect of the gene disruptions on fungal spread or mycotoxin content was detected, indicating that the ability of the fungus to manipulate the production of the gaseous plant hormones ethylene and H2S is dispensable for full virulence.

Pentahydroxyscirpene-Producing Strains, Formation In Planta, and Natural Occurrence.

Trichothecenes are a class of structurally diverse mycotoxins with more than 200 naturally occurring compounds. Previously, a new compound, pentahydroxyscirpene (PHS), was reported as a byproduct of a nivalenol producing Fusarium strain, IFA189. PHS contains a hydroxy group at C-8 instead of the keto group of type B trichothecenes. In this work, we demonstrate that IFA189 belongs to the species Fusarium kyushuense using molecular tools. Production of PHS in vitro was also observed for several isolates of other Fusarium species producing nivalenol. Furthermore, we report the formation of 4-acetyl-PHS by F. kyushuense on inoculated rice. Wheat ears of the variety Remus were infected with IFA189 and the in planta production of PHS was confirmed. Natural occurrence of PHS was verified in barley samples from the Czech Republic using a liquid chromatographic-tandem mass spectrometric method validated for this purpose. Toxicity of PHS to wheat ribosomes was evaluated with a coupled in vitro transcription and translation assay, which showed that PHS inhibits protein biosynthesis slightly less than nivalenol and deoxynivalenol.

Tracking by an optimal sequence of linear predictors.

We propose a learning approach to tracking explicitly minimizing the computational complexity of the tracking process subject to user-defined probability of failure (loss-of-lock) and precision. The tracker is formed by a Number of Sequences of Learned Linear Predictors (NoSLLiP). Robustness of NoSLLiP is achieved by modeling the object as a collection of local motion predictors--object motion is estimated by the outlier-tolerant RANSAC algorithm from local predictions. Efficiency of the NoSLLiP tracker stems from (i) the simplicity of the local predictors and (ii) from the fact that all design decisions--the number of local predictors used by the tracker, their computational complexity (i.e. the number of observations the prediction is based on), locations as well as the number of RANSAC iterations are all subject to the optimization (learning) process. All time-consuming operations are performed during the learning stage--tracking is reduced to only a few hundreds integer multiplications in each step. On PC with 1xK8 3200+, a predictor evaluation requires about 30 microseconds. The proposed approach is verified on publicly-available sequences with approximately 12000 frames with ground-truth. Experiments demonstrates, superiority in frame rates and robustness with respect to the SIFT detector, Lucas-Kanade tracker and other trackers.