Antioxidative defence mechanisms in two grapevine (Vitis vinifera L.) cultivars grown under boron excess in the irrigation water

The aim of this study was to investigate the impact of B excess on the antioxidative defence system in two grapevine cultivars differently sensitive to B, in order to evaluate in which way the different sensitivity was correlated to the activation of defence mechanisms. Two-year-old Vitis vinifera L. plants (‘Merlot’ and ‘Sangiovese’), grafted on the same rootstock, were subjected to B treatment as potted vines, and biochemical determinations were employed to evaluate the antioxidative response. Compared to ‘Sangiovese’, ‘Merlot’ showed a much higher B accumulation and both leaf and peroxidative damages. In B-treated ‘Sangiovese’ the activity of superoxide dismutase did not change compared to control leaves, whereas in ‘Merlot’ a dramatic decrease in the enzyme activity was observed. B increased reduced ascorbate pools in both cultivars, but ascorbate peroxidase activity was enhanced only in ‘Merlot’. In this latter cv. an enhancement of total phenols was also observed. ‘Merlot’ showed a lower ability to contrast B accumulation in leaves than ‘Sangiovese’ evidencing a higher oxidative stress. Even if defence mechanisms were generally activated in ‘Merlot’, they did not counteract efficiently metabolic damages likely due to the dramatic decrease in superoxide dismutase, the first enzyme involved in the detoxification of oxygen radicals

Lipid composition of pea (Pisum sativum L.) and maize (Zea mays L.) root plasma membrane and membrane-bound peroxidase and superoxide dismutase

Plasma membrane was isolated from roots of pea and maize plants and used to analyze POD and SOD isoforms, as well as lipid composition. Among lipids, phospholipids were the main lipid class, with phosphatidylcholine being the most abundant individual component in both pea and maize plasma membranes. Significant differences between the two plant species were found in the contents of cerebrosides, free sterols, and steryl glycosides. Most maize POD isoforms were with neutral and anionic pI values, but the opposite was observed in pea. While both anionic and cationic SOD isoforms were isolated from maize, only two anionic SOD isoforms were detected in pea

The Kinetic Effect of Some Wine Components on the Enzymatic Hydrolysis of β-glucan

Enzymatic preparations containing β-glucanases are utilised extensively in winemaking to facilitate the filtration of musts and wines coming from grapes affected by Botrytis cinerea, and to induce the release of mannoproteins and oligosaccharides from the cell walls of yeasts. The aim of the present work was to investigate the possible inhibitory effect of some wine components, and in particular of ethanol, on β-glucanase activities. For this purpose, the kinetic activity of a commercial enzymatic preparation containing β-glucanases was tested utilising both model solutions (buffer solution having a pH value similar to a must/wine with or without 13% v/v of ethanol added) and a red wine. When ethanol was added to the model solution, both the kinetic constant k and glucose production suffered a decrease of about 50% compared to the values detected in the absence of ethanol. A further loss of activity (about 87%) was found using red wine as reaction medium, suggesting, as already reported in the literature, that phenols could add their inhibitory effect to that of ethanol. The results obtained provide useful suggestions that can be adopted during winemaking. To promote the hydrolysis of the possible excess of β-glucan coming from grapes affected by Botritis cinerea, it would be more convenient to add β-glucanases before a significant amount of ethanol accumulates in the must/wine, while it is not possible to avoid the negative impact of ethanol or other possible inhibitors (phenols) to promote yeast degradation in wine left on lees

Foliar and root comparative metabolomics and phenolic profiling of Micro-Tom tomato (Solanum lycopersicum L.) plants associated with a gene expression analysis in response to short daily UV treatments

Tomato (Solanum lycopersicum L.) is globally recognised as a high-value crop both for commercial profit and nutritional benefits. In contrast to the extensive data regarding the changes in the metabolism of tomato fruit exposed to UV radiation, less is known about the foliar and root metabolome. Using an untargeted metabolomic approach through UHPLC-ESI-QTOF-MS analysis, we detected thousands of metabolites in the leaves (3000) and roots (2800) of Micro-Tom tomato plants exposed to 11 days of short daily UV radiation, applied only on the aboveground organs. Multivariate statistical analysis, such as OPLS-DA and volcano, were performed to allow a better understanding of the modifications caused by the treatment. Based on the unexpected modulation to the secondary metabolism, especially the phenylpropanoid pathway, of which compounds were down and up accumulated respectively in leaves and roots of treated plants, a phenolic profiling was carried out for both organs. The phenolic profile was associated with a gene expression analysis to check the transcription trend of genes involved in the UVR8 signalling pathway and the early steps of the phenolic biosynthesis. The retention of the modifications at metabolic and phenolic levels was also investigated 3 days after the UV treatment, showing a prolonged effect on the modulation once the UV treatment had ceased

A Kinetic Method to Identify the Optimum Temperature for β-glucanase Activity

Enzymatic preparations containing β-glucanases are largely utilised in winemaking to facilitate the filtration of musts and wines coming from grapes affected by Botrytis cinerea, and to induce the release of mannoproteins and oligosaccharides from the cell wall of yeasts. The aim of the present work was to investigate the effect of temperature on β-glucanase activity. For this purpose, the kinetic activity of a commercial enzymatic preparation containing β-glucanases was tested utilising a model solution (buffer solution of β-glucan extracted from alcoholic yeasts with a pH similar to a must/wine, with or without 13% ethanol added) at different temperatures ranging from 20 to 40°C. By an innovative procedure based on a kinetic model able to describe the time evolution of D-glucose accumulation – the final product of β-glucan hydrolysis – it was possible to relate the maximum rate of this process to temperature. The temperature at which the enzymatic activity showed the maximum value (Tmax) was close to 30°C, without any substantial variation following ethanol addition. Indeed, in spite of a remarkable reduction (of about 66%) in the catalytic activity shown by the commercial enzymatic preparation, no significant variation of the Tmax was observed, suggesting that the presence of an inhibitor such as ethanol in the reaction medium did not change the sensibility of the enzymes to temperature

Antifungal activity of azole compounds CPA18 and CPA109 against azole-susceptible and -resistant strains of Candida albicans

Objectives: In this study we investigated the in vitro fungistatic and fungicidal activities of CPA18 and CPA109, two azole compounds with original structural features, alone and in combination with fluconazole against fluconazole-susceptible and -resistant Candida albicans strains. Methods: Antifungal activities were measured by MIC evaluation and time–kill studies. Azole binding analysis was performed by UV-Vis spectroscopy. Hyphal growth inhibition and filipin and propidium iodide staining assays were used for morphological analysis. An analysis of membrane lipids was also performed to gauge alterations in membrane composition and integrity. Synergism was calculated using fractional inhibitory concentration indices (FICIs). Evaluation of cytotoxicity towards murine macrophages was performed to verify selective antifungal activity. Results: Even though their binding affinity to C. albicans Erg11p is comparable to that of fluconazole, CPA compounds are active against resistant strains of C. albicans with a mutation in ERG11 sequences and/or overexpressing the ABC transporter genes CDR1 and CDR2, which encode ATP-dependent efflux pumps. Moreover, CPA18 is fungistatic, even against the two resistant strains, and was found to be synergistic with fluconazole. Differently from fluconazole and other related azoles, CPA compounds induced marked changes in membrane permeability and dramatic alterations in membrane lipid composition. Conclusions: Our outcomes suggest that CPA compounds are able to overcome major mechanisms of resistance in C. albicans. Also, they are promising candidates for combination treatment that could reduce the toxicity caused by high fluconazole doses, particularly in immunocompromised patients

Cultivar-specific transcriptome prediction and annotation in Ficus carica L.

The availability of transcriptomic data sequence is a key step for functional genomics studies. Recently, a repertoire of predicted genes of a Japanese cultivar of fig (Ficus carica L.) was released. Because of the great phenotypic variability that can be found in this species, we decided to study another fig genotype, the Italian cv. Dottato, in order to perform comparative studies between the two cultivars and extend the pan genome of this species. We isolated, sequenced and assembled fig genomic DNA from young fruits of cv. Dottato. Then, putative gene sequences were predicted and annotated. Finally, a comparison was performed between cvs. Dottato and Horaishi predicted transcriptomes. Our data provide a resource (available at the Sequence Read Archive database under SRP109082) to be used for functional genomics of fig, in order to fill the gap of knowledge still existing in this species concerning plant development, defense and adaptation to the environment

Suppression of Phospholipase Dγs Confers Increased Aluminum Resistance in Arabidopsis thaliana

Aluminum (Al) toxicity is the major stress in acidic soil that comprises about 50% of the world's arable land. The complex molecular mechanisms of Al toxicity have yet to be fully determined. As a barrier to Al entrance, plant cell membranes play essential roles in plant interaction with Al, and lipid composition and membrane integrity change significantly under Al stress. Here, we show that phospholipase Dγs (PLDγs) are induced by Al stress and contribute to Al-induced membrane lipid alterations. RNAi suppression of PLDγ resulted in a decrease in both PLDγ1 and PLDγ2 expression and an increase in Al resistance. Genetic disruption of PLDγ1 also led to an increased tolerance to Al while knockout of PLDγ2 did not. Both RNAi-suppressed and pldγ1-1 mutants displayed better root growth than wild-type under Al stress conditions, and PLDγ1-deficient plants had less accumulation of callose, less oxidative damage, and less lipid peroxidation compared to wild-type plants. Most phospholipids and glycolipids were altered in response to Al treatment of wild-type plants, whereas fewer changes in lipids occurred in response to Al stress in PLDγ mutant lines. Our results suggest that PLDγs play a role in membrane lipid modulation under Al stress and that high activities of PLDγs negatively modulate plant tolerance to Al