Identification of Differently Regulated Proteins afterFusarium graminearum Infection of Emmer (Triticum dicoccum) at Several Grain Ripening Stages.

This study was conducted to improve the knowledge of molecular processes involved in the interaction between Fusarium graminearum and emmer in the course of grain ripening. Emmer plants were artificially inoculated with a F. graminearum spore suspension at anthesis. In the course of grain ripening from milk ripe to plant death stage, grains at four phenological growth stages were collected for analysis. The infection degree was evaluated based on the F. graminearum DNA content in emmer grain infolding tissues (glumes and rachis). For proteome analysis the albumin and globulin fractions of emmer grains, consisting of proteins with various functions related to the development and stress response, were analysed regarding the changes due to Fusarium infection by two-dimensional gel electrophoresis. Altogether, forty-three proteins affected by infection were identified by mass spectrometry. Enzymes detoxifying reactive oxygen species were regulated at all developmental stages. In the early stage of grain development, the abundance of proteins related to stress response, such as 2-Cys peroxiredoxin, a chitinase, a xylanase inhibitor and a spermidine synthase was increased. During later stage of grain development, the abundance of stress-related proteins, such as chitinases, heat shock proteins and an α-amylase inhibitor-like protein, decreased. During all ripening stages, but especially during medium milk stage (BBCH 75) and soft dough stage (BBCH 85), the abundance of proteins related to carbon metabolism, starch and protein biosynthesis as well as photosynthesis increased due to F. graminearum infection. At the plant death stage (BBCH 97) the abundance of only two proteins related to metabolism decreased.

Aflatoxin in Chili Peppers in Nigeria: Extent of Contamination and Control Using Atoxigenic Aspergillus flavus Genotypes as Biocontrol Agents.

Across sub-Saharan Africa, chili peppers are fundamental ingredients of many traditional dishes. However, chili peppers may contain unsafe aflatoxin concentrations produced by Aspergillus section Flavi fungi. Aflatoxin levels were determined in chili peppers from three states in Nigeria. A total of 70 samples were collected from farmers' stores and local markets. Over 25% of the samples contained unsafe aflatoxin concentrations. The chili peppers were associated with both aflatoxin producers and atoxigenic Aspergillus flavus genotypes. Efficacy of an atoxigenic biocontrol product, Aflasafe, registered in Nigeria for use on maize and groundnut, was tested for chili peppers grown in three states. Chili peppers treated with Aflasafe accumulated significantly less aflatoxins than nontreated chili peppers. The results suggest that Aflasafe is a valuable tool for the production of safe chili peppers. Use of Aflasafe in chili peppers could reduce human exposure to aflatoxins and increase chances to commercialize chili peppers in premium local and international markets. This is the first report of the efficacy of any atoxigenic biocontrol product for controlling aflatoxin in a spice crop.

Identification of regulated proteins in naked barley grains (Hordeum vulgare nudum) after Fusarium graminearum infection at different grain ripening stages.

We analyzed the effect of Fusarium graminearum infection on field-grown naked barley (Hordeum vulgare nudum). The ears were inoculated with F. graminearum spores during anthesis. In the course of ripening, grains in five phenological growth stages of naked barley from milk ripe to plant death were sampled. The albumin and globulin proteins of inoculated grains and untreated (control) grains were separated by two-dimensional gel electrophoresis. Forty-five spots composing of proteins that were changed in abundance due to F. graminearum infection were subsequently identified by mass spectrometry. Various proteins showing altered expression pattern after Fusarium infection were linked to stress response such as plant signal transduction pathways, fungal defense and oxidative burst. More proteins changed during early grain ripening stages than during later ripening stages. Protease inhibitors occurred at increased abundancy during milk ripe stage. A thaumatin-like protein accumulated at plant death stage. Proteins linked to nitrogen metabolism and protein biosynthesis were mainly reduced, whereas those linked to carbon metabolism were predominantly increased in infected grains. BIOLOGICAL SIGNIFICANCE: Fusarium graminearum infection can lead to significant contamination of grains with mycotoxins. With this 2D-based proteomics study we give an insight into plant­pathogen interactions between the non-model plant naked barley and the fungus F. graminearum during five stages of grain development. Over the multiple developmental stages we observed specific patterns of changes induced by the fungus: the primary plant metabolism and inhibition of fungal protease were predominantly affected during early grain development stages. During the entire grain development we found an induced accumulation of thaumatin-like proteins due to the fungal infection indicating their fundamental role for naked barley defense.

Determination of Ochratoxin A in Wheat and Maize by Solid Bar Microextraction with Liquid Chromatography and Fluorescence Detection.

Solid bar microextraction (SBME), followed by liquid chromatography with fluorescence detection (HPLC-FLD), for the quantification of ochratoxin A in wheat and maize was developed. Ground wheat and maize grains were extracted with acetonitrile-water-acetic acid (79:20:1, v/v/v), followed by defatting with cyclohexane, and subjected to SBME-LC-FLD analysis. SBME devices were constructed by packing 2 mg sorbent (C18) into porous polypropylene micro-tubes (2.5 cm length, 600 µm i.d., and 0.2 µm pore size). SBME devices were conditioned with methanol and placed into 5 mL stirred sample solutions for 70 min. After extraction, OTA was desorbed into 200 µL of methanol for 15 min, the solution was removed in vacuum, the residue was dissolved in 50 µL of methanol-water (1:1, v/v) and ochratoxin A content was determined by HPLC-FLD. Under optimized extraction conditions, the limit of detection of 0.9 µg·kg(-1) and 2.5 µg·kg(-1) and the precision of 3.4% and 5.0% over a concentration range of 1 to 100 µg·kg(-1) in wheat and maize flour, respectively, were obtained.