No abstract

This paper describes the first map, albeit partial, of toxigenic fungi re-isolated from grape berries collected in three out of the six most important Slovakia winemaking areas in two different periods of the harvest year 2008. Low temperatures and high relative humidity during July 2008 favoured the development of grape fungal diseases that cause rots such as Plasmopara, Uncinula, Botrytis, Metasphaeria, Elsinoë, and Saccharomycetes. In the analysed samples, the following genera of toxigenic fungi were identified in the range of 1-4%: Aspergillus, Alternaria, Cladosporium, Epicoccum, Fusarium, Penicillium, Rhizopus, Ulocladium, and Trichoderma Trichothecium, while the genera Aspergillus, Alternaria, Fusarium, and Penicillium were in the range 11-29%. A. niger, A. carbonarius, some strains of A. carbonarius-with 'crystals' and strains of A. uvarum-uniseriate were identified; these species are considered ochratoxigenic (able to produce variable amounts of toxins). In addition, a non-ochratoxigenic strain of A. ibericus and a Fusarium strain able to biosynthesize small amount of fumonisins, beauvericin, and enniatins were identified. P. expansum, able to produce citrinin, represents 29.7%, of the Penicillium genus together with P. verrucosum, P. glabrum, P. citrinum, and P. crustosum. An analysis for the identification and quantification of the main toxins: ochratoxin A, fumonisins, beauvericin, enniatins, and fusaproliferin was performed on grape samples; it was consistent with the results of the mycological analysis. Toxigenic fungi should be checked throughout the years and their occurrence compared with all environmental factors to avoid health risks.

Toxinogenic Fusarium species were identified on grape berries from Slovak vineyards, and their toxic metabolites were analysed by HPLC-MS/MS. F. subglutinans, F. oxysporum, F. proliferatum, F. semitectum, F. solani, F. subglutinans, and F. verticillioides were found with varying frequency. F. oxysporum and F. proliferatum, cultured in vitro on Czapek yeast autolysate agar and yeast extract sucrose agar, produced beauvericin, in the range from 3,265 to 13,400 μg/kg, and fusaproliferin in high concentration, ranging from 49,850 to 259,500 μg/kg. A maximum value of 2.24 μg/kg has been observed for beauvericin in dried grape berries. Fumonisin B1, and fumonisin B2 were also identified, and the observed levels ranged from 500 to 2,040 μg/kg. Over 2 years (namely 2008 and 2009) many other metabolites have been identified and analysed in grape berries, in particular: avenacein Y, apicidin, aurofusarin, chlamydosporol, 2-amino-14,16-dimethyloctadecan-3-ol, enniatin A, enniatin A1, enniatin B2, enniatin B3, and equisetin.

The secondary metabolite profiles of microfungi of the genus Penicillium isolated from samples of grape berries collected in two different phases during two vegetative seasons in Slovakia is described to assess the taxonomy. Three Slovak vine regions have been selected for this study, based on their climatic differences and national economic importance. Cultures of microfungi isolated from berries were incubated on different selective media for macro and micromorphology identification. The species Penicillium brevicompactum, Penicillium crustosum, Penicillium chrysogenum, Penicillium expansum, Penicillium palitans and Penicillium polonicum were identified according to growth and morphology. The related strains were found to produce a broad spectrum of fungal metabolites, including roquefortine C, chaetoglobosin A, penitrem A, cyclopeptin, cyclopenin, viridicatin, methylviridicatin, verrucofortine, secalonic acid D, cyclopiazonic acid, fumigaclavine and mycophenolic acid. Chemotaxonomy was performed using high-performance liquid chromatography (HPLC) and mass spectrometry (MS). Dried grape berries were also analyzed allowing to assess the presence of patulin, roquefortine C and penicillic acid; this last one has been identified in dried berries but not in vitro.

Recently, beauvericin (BEA) has been recognized as an important toxic compound synthesized by several Fusarium strains, infecting maize, wheat, and rice, worldwide. The effects of BEA on mammalian cells have been studied; however, its effects on the function of host plant cells are largely unknown. The purpose of our work was to assess whether BEA can affect the root and leaf cells of wheat cultivar (cv.) 'Arina' seedlings, using a cytotoxicity assay and fluorescence microscopy. Toxigenicity during wheat germination was higher in BEA-treated wheat seedlings than in non-treated seedlings (control). Leaf primordial, situated at the base and the tips of treated leaves, were more affected by BEA compared to the control when assayed in medium for cell viability measured by luminescent equipment. BEA-Treated plant cells secrete adenosine triphosphate (ATP) to the extracellular matrix and invoke more luminescence by luciferase than the non-treated seedlings. Our results were confirmed by fluorescence microscopy following '4',6-diamidino-2-phenylindole' (DAPI) staining and by confocal microscopy. In addition, the bioluminescent protein luciferase was observed in the intracellular space indicating presence of ATP. The incidence of nuclear fragmentation increased significantly in cells of seedlings treated with BEA at 40 microM concentration implying that the intracellular phytotoxin BEA plays an important role, possibly as a mediator in cell-death signalling.