Morphometric and molecular analysis of a pink-berried mutant within the population of grape cultivar 'Plavac mali'

This study reports characteristics of pink-berried mutants found in the clone population of the Croatian red wine cultivar 'Plavac mali' on the basis of comparative ampelographic and DNA marker (SSR and Gret1) analysis. The pink-berried accession, also called 'Plavac mali sivi' (Croatian: sivi = English: grey or French: gris), along with the other 58 'Plavac mali' accessions of standard blue-black berry skin color, has been characterized for the first time using OIV descriptors and molecular markers. Using a set of 9 simple sequence repeat (SSR) markers, an identical SSR profile for all the analyzed accessions was revealed, indicating their monozygotic status. The analysis of Gret1 insertion within the VvMYBA1 locus revealed no DNA polymorphism responsible for the pink-berried phenotype. Surface color of the berry skin was measured with the CIELab technique using a reflectance spectrophotometer at full ripeness. The results of colorimetric variables (L*, a* and b*) suggest a significantly lower accumulation of anthocyanins in the pink-berried accession compared to the standard blue-black berries. The pink-berried accession shares all assessed morphological and genetic traits of 'Plavac mali', with the only difference being the color of the berry skin. This suggests that the pinkberried genotype is the result of a spontaneous mutation of a standard 'Plavac mali' genotype

Analysis of phyllometric parameters efficiency in discrimination of Croatian native V. vinifera cultivars

The aims of this study were to achieve correct cultivar classification of leaf samples by using phyllometric parameters to determine the performance of parameters applied as the discriminant criteria and to determine the minimal number of leaf parameters needed to accurately classify leaf samples within cultivars. Seventy-nine phyllometric parameters were measured/calculated on 360 leaf samples from eleven grapevine cultivars, gathered during several years, from different growing conditions and with some differences in the sampling methods applied. Stepwise discriminant analysis was used to rank the phyllometric parameters according to their efficiency in the discrimination of cultivars. A series of discriminant analyses was performed with successive introduction of new parameters as discriminatory elements until 100 % correct classification of leaf samples into the correct cultivar was achieved. This was achieved using only the seven highest ranking phyllometric parameters from the stepwise discriminant analysis. Additionally, canonical discriminant analysis was performed to evaluate the differences between cultivars in the same parameters. This study represents a model for analysis of the efficiency of different ampelometric parameters for discrimination of V. vinifera cultivars.

Differences in chemical composition of 'Plavac mali' grape berries

Evaluation of differences in yield and quality of grapevine is often based on the random samples of berries harvested on the same date. Due to differences in ripeness of berries from the same cluster, and among berries from different clusters of the same vine, the determined differences among examined treatments (clones, agricultural practices etc.) based on random berry samples may be over- or underestimated. The aims of this study were to determine: (1) differences among three 'Plavac mali' clones in the proportion of berries of different density classes (sugar concentration level) using the flotation method; (2) differences in berry weight and chemical composition among density classes of the same clone, and (3) differences between clones of the same density class. Significant differences were determined for all observed characteristics, among different berries density classes and clones. The results obtained indicate that berry samples used to compare different clones should be, to the greatest possible extent, at the same level of ripeness, as this significantly effects chemical composition. This can be achieved using the density separation of berries with the simple flotation method presented in this study. The presented results could be useful in the future improvement of clonal selection methodology as well as in improvement of sampling strategy for other types of researches

Cryopreservation and cryotherapy of grapevine (Vitis vinifera L.)

This study aimed at testing the efficiency of a droplet-vitrification cryopreservation protocol in eliminating selected grapevine viruses. The cryopreservation protocol led to approximately 50 % recovery with cultivar 'Portan' and five international cultivars tested, but very low recovery was noted with Croatian cultivars. GFLV and GLRaV-3, two (economically important grapevine) viruses were eliminated in a high percentage (up to 100 %) of plants regenerated from cryopreserved shoot tips. Virus sanitation was observed as well in samples before liquid nitrogen exposure. Genetic stability of plants regenerated after cryopreservation was studied using AFLP markers. Polymorphic fragments were observed in non-cryopreserved and cryopreserved samples treated with PVS2 solution, the number of which increased with increasing durations of exposure to PVS2 solution

Ampelographic and genetic characterization of Croatian grapevine varieties

Before the Phylloxera (Daktulosphaira vitifoliae) crisis in Croatia more than 400 grape varieties were under cultivation. Today their number is drastically decreased. Recently, many efforts in the preservation of grapevine biodiversity were undertaken: detailed inventory of varieties in different wine-growing regions, establishing of national and regional collections and their systematic ampelographic and genetic characterisation. This paper shows results of multiannual ampelographic and genetic research concerning Croatian grapevines considered as autochthonous varieties. After identification, many synonyms and homonyms were detected and unique genotypes were selected. Basic data of them are shown: description (according to OIV descriptors) and genetic profiles (9 SSR loci), growing area and status of vulnerability. Also, genetic relationship based on the shared allele distance is computed from SSR data. Results show certain level of similarity among varieties, and classify Croatia as important gene-pool in Europe. Some additional accessions which were detected and introduced into collections recently are still being investigated and characterized so that the here presented list of Croatian native grapevine cultivars is not final

Energy at Origins: Favorable Thermodynamics of Biosynthetic Reactions in the Last Universal Common Ancestor (LUCA)

Though all theories for the origin of life require a source of energy to promote primordial chemical reactions, the nature of energy that drove the emergence of metabolism at origins is still debated. We reasoned that evidence for the nature of energy at origins should be preserved in the biochemical reactions of life itself, whereby changes in free energy, ΔG, which determine whether a reaction can go forward or not, should help specify the source. By calculating values of ΔG across the conserved and universal core of 402 individual reactions that synthesize amino acids, nucleotides and cofactors from H2, CO2, NH3, H2S and phosphate in modern cells, we find that 95-97% of these reactions are exergonic (ΔG ≤ 0 kJ⋅mol-1) at pH 7-10 and 80-100°C under nonequilibrium conditions with H2 replacing biochemical reductants. While 23% of the core's reactions involve ATP hydrolysis, 77% are ATP-independent, thermodynamically driven by ΔG of reactions involving carbon bonds. We identified 174 reactions that are exergonic by -20 to -300 kJ⋅mol-1 at pH 9 and 80°C and that fall into ten reaction types: six pterin dependent alkyl or acyl transfers, ten S-adenosylmethionine dependent alkyl transfers, four acyl phosphate hydrolyses, 14 thioester hydrolyses, 30 decarboxylations, 35 ring closure reactions, 31 aromatic ring formations, and 44 carbon reductions by reduced nicotinamide, flavins, ferredoxin, or formate. The 402 reactions of the biosynthetic core trace to the last universal common ancestor (LUCA), and reveal that synthesis of LUCA's chemical constituents required no external energy inputs such as electric discharge, UV-light or phosphide minerals. The biosynthetic reactions of LUCA uncover a natural thermodynamic tendency of metabolism to unfold from energy released by reactions of H2, CO2, NH3, H2S, and phosphate

Molecular characterization of old local grapevine varieties from South East European countries

South East European (SEE) viticulture partially relies on native grapevine varieties, previously scarcely described. In order to characterize old local grapevine varieties and assess the level of synonymy and genetic diversity from SEE countries, we described and genotyped 122 accessions from Albania, Federation of Bosnia and Herzegovina (B&H), Croatia, Macedonia, Moldova, Montenegro, Republika Srpska (Bosnia and Herzegovina) and Romania on nine most commonly used microsatellite loci. As a result of the study a total of 86 different genotypes were identified. All loci were very polymorphic and a total of 96 alleles were detected, ranging from 8 to 14 alleles per locus, with an average allele number of 10.67. Overall observed heterozygosity was 0.759 and slightly lower than expected (0.789) while gene diversity per locus varied between 0.600 (VVMD27) and 0.906 (VVMD28). Eleven cases of synonymy and three of homonymy have been recorded for samples harvested from different countries. Cultivars with identical genotypes were mostly detected between neighboring countries. No clear differentiation between countries was detected although several specific alleles were detected. The integration of the obtained genetic data with ampelographic ones is very important for accurate identification of the SEE cultivars and provides a significant tool in cultivar preservation and utilization.

The European Vitis Database (www.eu-vitis.de) – a technical innovation through an online uploading and interactive modification system

The objective of the European Vitis Database is to safeguard and enhance germplasm by monitoring its preservation. Two issues are strongly related to that purpose: (1) participation of collections covering almost all grape biodiversity and (2) assessment of accessions trueness to type. In the scope of the European project GrapeGen06 efforts have been made towards both objectives. The 35 participating grape germplasm repositories are found between the Iberian Peninsula and Transcaucasia, thus covering a broad range of grape diversity. Altogether they maintain 32,410 accessions. However with respect to biodiversity, gaps are still evident and further collections need to be included and trueness to type assessment absolutely needs to be pursued to organize duplication of endangered genotypes. Within the GrapeGen06 project focus was laid on the establishment of a database conferring the collection holders a high degree of responsibility and independence. Hence for the first time in a European Central Crop Database an on-line uploading application and an interactive modification system for data administration was implemented. These innovations disburden the database manager and offer the curators of collections more flexibility. Prerequisites for data import, descriptors applied, access levels, database contents, uploading, export and search functions are described

Description of the Vitis vinifera L. phenotypic variability in eno-carpological traits by a Euro-Asiatic collaborative network among ampelographic collections

The grapevine intra-specific variability captured an increasing interest during the last decades, as demonstrated by the number of recently funded European projects focused on the grapevine biodiversity preservation. However, nowadays, crop plants are mainly characterized by genotyping methods. The present work summarizes the phenotype data collected among 20 ampelographic collections spread in 15 countries, covering mostly of the viticultural areas in the Euro-Asiatic range: from Portugal to Armenia and from Cyprus to Luxembourg. Together with agro-climatic characterization of the experimental site, in two years, about 2400 accessions were described, following a common experimental protocol mainly focused on the carpological and oenological traits, obtaining a general overview of the distribution of the considered phenotypic traits in the cultivated Vitis vinifera species. The most replicated cultivars were selected and, for the subset of these reference cultivars, their behavior in the different environmental conditions over sites and years was described by ANOVA methods