This paper represents the results of screening a diversity of fungal endophytes associated with Vitis vinifera leaves and canes in the Czech Republic. The characterization of strains is based on morphological and phylogenetic analyses of ITS, EF1α and TUB2 sequence data. Our strain selection covers 16 species and seven orders belonging to Ascomycota and Basidiomycota. Together with ubiquitous fungi, we report on several poorly known plant-associated fungi, Angustimassarina quercicola (= A. coryli, a synonym proposed in this study) and Pleurophoma pleurospora. Other species, such as Didymella negriana, D. variabilis, Neosetophoma sp. (species identical or sister to N. rosae), Phragmocamarosporium qujingensis and Sporocadus rosigena, have so far been little known and rarely found, but are frequent on V. vinifera in different parts of the world and obviously belong to a microbiota with a strong preference for this plant. Detailed taxonomical identification allowed us to identify species with apparent stable associations with V. vinifera, for which further interactions with V. vinifera can be expected. Our study is the first to focus on V. vinifera endophytes in Central Europe and expands the knowledge about their taxonomy, ecology and geography.
- MeSH
- Basidiomycota * MeSH
- Endophytes genetics MeSH
- Phylogeny MeSH
- Fungi MeSH
- Vitis * microbiology MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Czech Republic MeSH
We investigated ochratoxin A (OTA) contamination in raisin samples purchased from Slovak markets and determined the diversity of black-spored aspergilli as potential OTA and fumonisin (FB1 and FB2) producers. The taxonomic identification was performed using sequences of the nuclear ITS1-5.8s-ITS2 region, the calmodulin and beta-tubulin genes. We obtained 239 isolates from eight fungal genera, of which 197 belonged to Aspergillus (82%) and 42 strains (18%) to other fungal genera. OTA contamination was evidenced in 75% of the samples and its level ranged from 0.8 to 10.6 µg/kg. The combination of all three markers used enabled unambiguous identification of A. carbonarius, A. luchuensis, A. niger, A. tubingensis and A. welwitschiae. The dominant coloniser, simultaneously having the highest within-species diversity isolated from our raisin samples, was A. tubingensis. Out of all analysed strains, only A. carbonarius was found to produce OTA, but in relatively high quantity (2477-4382 µg/kg). The production of FB1 and FB2 was evidenced in A. niger strains only.
- MeSH
- Aspergillus genetics metabolism MeSH
- Fumonisins metabolism toxicity MeSH
- Phylogeny MeSH
- Genetic Variation * MeSH
- Risk Assessment MeSH
- Food Preservation MeSH
- Ochratoxins metabolism toxicity MeSH
- Fruit microbiology MeSH
- Food Microbiology MeSH
- Vitis microbiology MeSH
- Desiccation MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
BACKGROUND: The study was focused on Zn, Cu and Pb transfer in the system of soil-grape-must-juice-wine in a Chardonnay grape variety from Ukrainian vine growing regions. The analyses of soil, grape, must, pomace, juice and wine were done at the study plot in the south-west of Crimea. RESULTS: Commercial white wines of Chardonnay from different vine growing regions in Ukraine were analysed for trace metals content. Results revealed that trace elements transfer was related to diverse Zn, Cu and Pb sources, trace metals bioavailability, their speciation and complexes during the wine making processes. The analysed commercial wines had lower Cu, Zn and Pb concentrations than wine from the Inkerman study plot. CONCLUSION: Trace metals concentrations were comparable to those in European wines and lower than limits recommended by International Organization of Vine and Wine. The tentative relationship between wine and soil was found for Zn at the study plot. The method can be used to describe the relationship between the soil and wine in other study areas. © 2017 Society of Chemical Industry.
- MeSH
- Soil Pollutants analysis MeSH
- Food Handling MeSH
- Copper analysis MeSH
- Lead analysis MeSH
- Fruit chemistry growth & development MeSH
- Fruit and Vegetable Juices analysis MeSH
- Soil chemistry MeSH
- Trace Elements analysis MeSH
- Wine analysis MeSH
- Vitis chemistry growth & development MeSH
- Zinc analysis MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Ukraine MeSH
Two metal(loid) contaminated agricultural soils were amended with grape stalk (wine production by-product)-derived biochar as well as its pre-pyrolysed origin material, to investigate their geochemical impacts on As, Cr, Cu and Zn. Detailed physico-chemical evaluation combined with a column leaching test determined the retention of metal(loid)s from soil solution by each amendments. A pot experiment measured metal(loid)s in soil pore water and their uptake to ryegrass when the amendments were mixed into soils at 1 and 5% (w/w). Total Cr and Zn concentrations were reduced furthest in column leachates by the addition of raw material and biochar respectively, compared to the untreated soil; Cr(III) was the predominant specie initially due to rapid acidification of leachates and organic complexation resulting from raw material addition. Loadings of metal(loid)s to the amendments recovered from the post-leached columns were in the order Cu » Zn > Cr ≈ As. In the pot test ryegrass Cr uptake was initiated by the addition of both amendments, compared to the untreated soil, whereas only biochar addition resulted in significant increases in Zn uptake, explained by its significant enhancement of ryegrass biomass yield, especially at 5% dosage; raw material addition significantly decreased biomass yields. Inconsistent relationships between pore water parameters and ryegrass uptake were common to both soils investigated. Therefore, whilst both amendments modified soil metal(loid) geochemistry, their effects differed fundamentally; in environmental risk management terms these results highlight the need to investigate the detailed geochemical response of contaminated soils to diverse organic amendment additions.
Comprehensive next generation sequencing virus detection was used to detect the whole spectrum of viruses and viroids in selected grapevines from the Czech Republic. The novel NGS approach was based on sequencing libraries of small RNA isolated from grapevine vascular tissues. Eight previously partially-characterized grapevines of diverse varieties were selected and subjected to analysis: Chardonnay, Laurot, Guzal Kara, and rootstock Kober 125AA from the Moravia wine-producing region; plus Müller-Thurgau and Pinot Noir from the Bohemia wine-producing region, both in the Czech Republic. Using next generation sequencing of small RNA, the presence of 8 viruses and 2 viroids were detected in a set of eight grapevines; therefore, confirming the high effectiveness of the technique in plant virology and producing results supporting previous data on multiple infected grapevines in Czech vineyards. Among the pathogens detected, the Grapevine rupestris vein feathering virus and Grapevine yellow speckle viroid 1 were recorded in the Czech Republic for the first time.
- MeSH
- DNA, Viral chemistry MeSH
- Farms MeSH
- Plant Viruses isolation & purification MeSH
- Sequence Analysis, DNA methods MeSH
- Viroids isolation & purification MeSH
- Vitis virology MeSH
- High-Throughput Nucleotide Sequencing methods MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Czech Republic MeSH
Many different yeast species have been isolated from grapes and musts worldwide. The diversity and frequency of yeasts depend on a number of factors such as the grape variety, the physical damage of the grapes, the weather conditions and the chemical composition of must. A total of 366 isolates were associated with the three grape cultivars: Blue Frankish, Green Veltliner and Sauvignon blanc over four consecutive years. Yeast cultures were isolated from the grapes and from the fermenting musts after the first and seventh days. The ascomycetous yeasts of the genera Aureobasidium, Candida, Hanseniaspora, Metschnikowia, Pichia, Saccharomyces and Saccharomycopsis together with basidiomycetous yeasts of the genera Cryptococcus, Dioszegia, Filobasidium, Rhodotorula and Sporidiobolus were associated with the three grape varieties. Hanseniaspora uvarum, Metschnikowia pulcherrima, Pichia kluyveri, Pichia kudriavzevii and Sporidiobolus pararoseus were found on the berries in significant amounts. P. kluyveri and P. kudriavzevii were more associated with the damaged grapes, whereas Sp. pararoseus with intact ones. H. uvarum and M. pulcherrima were present on both types of grapes almost equally. The yeast composition and quantitative representation of yeast species varied over the grape varieties and the years examined. Although the basidiomycetous species formed a significant proportion of the yeast population in some individual grape variety/year combinations, the ascomycetous species were dominant.
- MeSH
- Biodiversity * MeSH
- DNA, Fungal chemistry genetics MeSH
- RNA, Fungal genetics MeSH
- Phylogeny MeSH
- Genes, rRNA MeSH
- Fungi classification genetics isolation & purification MeSH
- DNA, Ribosomal chemistry genetics MeSH
- RNA, Ribosomal genetics MeSH
- Sequence Analysis, DNA MeSH
- Wine microbiology MeSH
- Vitis microbiology MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Slovakia MeSH
The diversity of yeasts and yeast microorganisms on grapes and, consequently, in musts may be strongly influenced by using Cu2+ products. The effect of Cu2+ concentration on the growth of native yeast strains isolated from grapes and grape must of three grape varieties was studied. The growth of H. uvarum, P. kluyveri, P. kudriavzevii, C. californica, F. elegans, R. bacarum and Sc. crataegensis was inhibited in the presence of a minimum concentration of 0.5 mM Cu2+. The growth of S. cerevisiae, Cr. magnus, R. glutinis and Sp. pararoseus depended on the strain tested. The strains of A. pullulans, M. pulcherrima, C. oleophila, P. terricola, R. nothofagi and R. minuta were growing at increasing Cu2+ concentrations but their growth was reduced by prolongation of the lag phase compared with their growth in the absence of Cu2+. The growth reduction of ethanol-tolerant yeast strains at higher concentration of Cu ions shows the effect of the prolonged action of wine fermentation. Autochthonous yeast strains resistant to higher Cu2+ concentrations (C. oleophila, M. pulcherrima, R. minuta) appear to be an appropriate solution to reduction of Cu residues in the fermenting must.
There is relatively little information concerning long-term alterations in DNA methylation following exposure of plants to environmental stress. As little is known about the ratio of non-heritable changes in DNA methylation and mitotically-inherited methylation changes, dynamics and reversibility of the DNA methylation states were investigated in grapevine plants (Vitis vinifera) stressed by in vitro cultivation. It was observed that significant part of induced epigenetic changes could be repeatedly established by exposure to particular planting and stress conditions. However, once stress conditions were discontinued, many methylation changes gradually reverted and plants returned to epigenetic states similar to those of maternal plants. In fact, in the period of one to three years after in vitro cultivation it was difficult to distinguish the epigenetic states of somaclones and maternal plants. Forty percent of the observed epigenetic changes disappeared within a year subsequent to termination of stress conditions ending and these probably reflect changes caused by transient and reversible stress-responsive acclimation mechanisms. However, sixty percent of DNA methylation diversity remained after 1 year and probably represents mitotically-inherited epimutations. Sequencing of regions remaining variable between maternal and regenerant plants revealed that 29.3% of sequences corresponded to non-coding regions of grapevine genome. Eight sequences (19.5%) corresponded to previously identified genes and the remaining ones (51.2%) were annotated as "hypothetical proteins" based on their similarity to genes described in other species, including genes likely to undergo methylation changes following exposure to stress (V. vinifera gypsy-type retrotransposon Gret1, auxin-responsive transcription factor 6-like, SAM-dependent carboxyl methyltransferase).
- MeSH
- DNA, Plant analysis isolation & purification MeSH
- Electrophoresis, Capillary MeSH
- Epigenesis, Genetic MeSH
- Stress, Physiological * MeSH
- DNA Methylation * MeSH
- Plant Cells metabolism MeSH
- Sequence Analysis, DNA MeSH
- Temperature MeSH
- Vitis genetics growth & development MeSH
- Chromatography, High Pressure Liquid MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Grapevine Syrah virus-1 (GSyV-1) was identified by small-RNA deep sequencing in Slovak grapevine co-infected by several other viruses. The RT-PCR assays developed in this work substantially improved the virus detection and allowed the identification of GSyV-1 in tested grapevine samples from Slovakia and the Czech Republic at an unexpectedly high rate (ca. 30 %). Subsequently, complete genome sequences of 3 GSyV-1 isolates (2 Slovak and 1 Czech) were determined by Sanger sequencing, showing a typical marafivirus genome organization. Analyses of complete genome sequences showed a higher intra-group diversity among these 3 central European GSyV-1 isolates (differences reaching 7.1 % at the nucleotide level) in comparison to 3 previously characterized North American isolates (only 1.2 % intra-group divergence). A substantially higher divergence among central European isolates and their clustering into two major phylogenetic groups was further confirmed by the partial genome analysis of additional 26 isolates. The CP-centered study did not support the geography-based clustering among central European and American isolates. Nevertheless, the sequence data of the highly variable 5'-proximal portion of the genome obtained for few additional isolates from Slovakia and Czech Republic showed the presence of both, "European-" and "north American-like", GSyV-1 isolates in the analyzed grapevine samples.
- MeSH
- Phylogeny MeSH
- Genetic Variation MeSH
- Genome, Viral * MeSH
- Molecular Sequence Data MeSH
- Gene Order MeSH
- RNA, Viral genetics MeSH
- Sequence Analysis, DNA * MeSH
- Sequence Homology, Nucleic Acid MeSH
- Cluster Analysis MeSH
- Tymoviridae classification genetics isolation & purification MeSH
- Vitis virology MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Czech Republic MeSH
- Slovakia MeSH
Analysis of complete genome sequences of three Slovak isolates of grapevine Pinot gris virus (GPGV) showed their low heterogeneity (reaching 1.7 %) and a close relationship to the Italian NC_015782 isolate (4.2-4.5 % divergence). Comparison of Slovak and Italian isolates revealed an unusual accumulation of 21 indel mutations in ORF1, resulting in a localized high divergence in the encoded amino acid sequences. An elevated divergence in the 5' extremity of the GPGV genomes is suggestive of a recombination between Slovak isolates and grapevine berry inner necrosis virus. RT-PCR allowed the frequent detection of closely related GPGV isolates in grapevines from Slovakia and the Czech Republic.
- MeSH
- Flexiviridae classification genetics isolation & purification MeSH
- Phylogeny MeSH
- Genetic Variation * MeSH
- Genome, Viral MeSH
- Molecular Sequence Data MeSH
- Plant Diseases virology MeSH
- Open Reading Frames MeSH
- Vitis virology MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Czech Republic MeSH
- Slovakia MeSH
