Ahmed Mliki

Abscisic acid signals reorientation of polyamine metabolism to orchestrate stress responses via the polyamine exodus pathway in grapevine.

Polyamines (PAs) have been suggested to be implicated in plant responses to abiotic and biotic stress. Grapevine is a model perennial plant species whose cultivars respond differently to osmotic stress. In this study, we used two cultivars, one sensitive (S) and one tolerant (T) to drought. In adult vines subjected to drought under greenhouse conditions, total PAs were significantly lower in the control T- and higher in the control S-genotype and significantly increased or decreased, respectively, post-treatment. Soluble Put and Spd exhibited the greatest increase on d 8 post-treatment in the T- but not in the S-genotype, which accumulated soluble Spm. Abscisic acid (ABA) was differentially accumulated in T- and S-genotypes under drought conditions, and activated the PA biosynthetic pathway, which in turn was correlated with the differential increases in PA titers. In parallel, polyamine oxidases (PAOs) increased primarily in the S-genotype. ABA at least partially induced PA accumulation and exodus into the apoplast, where they were oxidized by the apoplastic amine oxidases (AOs), producing H2O2, which signaled secondary stress responses. The results here show that the ABA signaling pathway integrates PAs and AOs to regulate the generation of H2O2, which signals further stress responses or the PCD syndrome.

Water stress induced changes in the leaf lipid composition of four grapevine genotypes with different drought tolerance

To dissect differences in both lipid accumulation and composition and the role of these modifications during drought stress, four grapevine cultivars exhibiting differential tolerance to drought were subjected to water shortage. Tolerant cultivars, Kahli Kerkennah and Cardinal, exhibited higher leaf water potential (Ψw), and lower lipid peroxidation compared to the sensitive cultivars Guelb Sardouk and Superior Seedless during stress. Total lipid amounts increased during stress only in the leaves of the tolerant cultivars. Drought induced increases in the ratios digalactosyldiacylglycerol/monogalactosyldiacylglycerol and phosphatidylcholine/phoshatidylethanolamine of almost all the drought stressed cultivars. Moreover, the overall analysis of the composition of fatty acids revealed that a linolenic acid was prevalent in grapevine and the unsaturation level of lipids increased under water stress in all the cultivars. Specific adjustments in the lipid composition during stress could compromise stress tolerance.

Genetic diversity in melon (Cucumis melo L.): Anevaluation of African germplasm

The genetic diversity among 126 exotic (108) andreference array (RA) melon (Cucumismelo L.) accessions (18) was assessed byvariation at 49 random amplified polymorphic DNA marker bands(putative loci) using 29 10-mer primers. Africanaccessions of unknown melon market classes were compared to the RAaccessions from a broad range of C.melo subsp. melo groups(Cantalupensis, Conomon, Inodorus and Flexuosus). Althoughdifferences in groupings occurred after multidimensional scaling andcluster analysis, both analyses placed African accessions into twogroups, which were separate from RA groupings. One African group of33 accessions containing accessions from Zimbabwe (5),Zambia (24), Mali (1), one of two Senegalaccessions and two of three South African accessions examined. Thesecond group, which consisted of 67 accessions containing collectionsfrom Egypt (40), Tunisia (6), Libya(13), Morocco (1), Algeria (2),Ethiopia (1), Niger (1), Sierra Leone(1), S. Africa (1), Zambia (1) andZimbabwe (1). Depending on the multivariate analysistechnique employed, accessions from Kenya, Senegal and Ghana formedeither unique groupings or were grouped with accessions(Cantalupensis) from the RA. Both analyses indicate thatthe genetic differences inherent between the African gene pools isassociated with the geographic proximity of African countries(northern vs. central-southern Africa) in thegermplasm array examined. Moreover, these data indicate that thegenetic diversity of U.S. and European commercial RA germplasm(Cantalupensis and Inodorus) could be enhanced by theintroduction of genetic variation from African accessions, and thatit would be advantageous to acquire more accessions from thisgeographically and ecologically varied region to ensure the retentionof existing genetic diversity.

Greater Effectiveness of ε-Viniferin in Red Wine Than Its Monomer Resveratrol for Inhibiting Vascular Smooth Muscle Cell Proliferation and Migration

Resveratrol is a strong candidate for explaining an irreversible correlation between red wine consumption and coronary heart disease. The present study examined the effect of ε-viniferin, a dehydrodimer of resveratrol, on vascular smooth muscle cells (VSMCs), because ε-viniferin functions are poorly understood in spite of its comparable content to resveratrol in red wines and grapes. Both ε-viniferin and resveratrol inhibited platelet-derived growth factor-induced cell proliferation, migration, and reactive oxygen species (ROS) production, in addition to inducing nitric oxide generation. ε-Viniferin was more effective than resveratrol in these effects, except for inhibiting ROS production. The compounds also increased the expression of the antioxidant enzyme, hemeoxygenase-1, via transcription factor Nrf2. The phosphatidylinositol 3-kinase-Akt pathway was implicated in resveratrol-dependent nuclear Nrf2 accumulation, whereas extracellular signal-regulated kinase and p38 were involved in ε-viniferin-induced Nrf2 accumulation. These data suggest that ε-viniferin may function more effectively than resveratrol in different mechanisms and cooperatively with resveratrol in preventing atherosclerosis.

Thiamine modulates metabolism of the phenylpropanoid pathway leading to enhanced resistance to Plasmopara viticola in grapevine

BackgroundPreviously, we have reported the ability of thiamine (vitamin B1) to induce resistance against Plasmopara viticola in a susceptible grapevine cv. Chardonnay. However, mechanisms underlying vitamins, especially, thiamine-induced disease resistance in grapevine are still largely unknown. Here, we assessed whether thiamine could modulate phenylpropanoid pathway-derived phytoalexins in grapevine plants, as well as, the role of such secondary metabolites in thiamine-induced resistance process to P. viticola.ResultsOur data show that thiamine treatment elicited the expression of phenylpropanoid pathway genes in grapevine plants. The expression of these genes correlated with an accumulation of stilbenes, phenolic compounds, flavonoids and lignin. Furthermore, the total anti-oxidant potential of thiamine-treaded plants was increased by 3.5-fold higher level as compared with untreated-control plants. Four phenolic compounds are responsible of 97% of the total anti-oxidant potential of thiamine-treated plants. Among these compounds, is the caftaric acid, belonging to the hydroxy-cinnamic acids family. This element contributed, by its own, by 20% of this total anti-oxidant potential. Epifluorescence microscopy analysis revealed a concomitant presence of unbranched-altered P. viticola mycelia and stilbenes production in the leaf mesophyll of thiamine-treated inoculated plants, suggesting that stilbenes are an important component of thiamine-induced resistance in grapevine.ConclusionThis work is the first to show the role of thiamine, as a vitamin, in the modulation of grapevine plant secondary metabolism contributing to an enhanced resistance to P. viticola, the most destructive fungal disease in vineyards.

Identification et caractérisation d'un gène de réponse à la déshydratation « rd22 » chez la vigne (Vitis vinifera L.)

To identify and isolate genes related to abiotic stresses (salinity and drought) tolerance in grapevine, a candidate gene approach was developed and allowed isolating a full-length cDNA of rd22 gene from the Cabernet Sauvignon variety. The latter, named Vvrd22, is a dehydration-responsive gene that is usually induced by the application of exogenous ABA. Details of the physicochemical parameters and structural properties (molecular mass, secondary structure, conserved domains and motives, putative post-translational modification sites...) of the encoded protein have also been elucidated. The expression study of Vvrd22 was carried out at the berry growth stages and at the level of plant organs and tissues as well as under both drought and salt stresses. The results showed that Vvrd22 is constitutively expressed at a low level in all analyzed tissues. Moreover, salt stress induced Vvrd22 expression, particularly for the tolerant variety (Razegui), contrary to the sensitive one (Syrah), which did not display any expression variation during the stress, which means that Vvrd22 is involved in salt stress response and that its expression level depends on regulatory mechanisms that are efficient only for the tolerant variety. On the other hand, under drought stress, Vvrd22 is induced in an identical manner for both tolerant and sensitive varieties. In addition, stress signal molecules such as ABA (lonely applied or in combination with sucrose) induced Vvrd22 expression, even at a low level. A minimal knowledge about the role and the functionality of this gene is necessary and constitutes a prerequisite condition before starting and including Vvrd22 in any program of improvement of grapevines abiotic stress tolerance.

Isolation and expression analysis of salt induced genes from contrasting grapevine (Vitis vinifera L.) cultivars.

Salt stress adversely affects the growth of grapevine plants. In order to understand the molecular basis of salt stress response in grapevine plants, suppression subtractive hybridization (SSH) and microarray based screening approaches were combined. Two leaf-specific subtractive cDNA libraries were constructed from grapevine plants subjected to a moderate, incremental salt stress treatment. SSH were performed 6h and 24h after NaCl peaked at 100mM using cDNAs prepared from leaves of a salt tolerant cultivar (Razegui) as testers and cDNAs from unstressed leaves as drivers. Then, a pre-screened subset of cDNA clones from these SSH libraries were used to construct a Vitis vinifera cDNA array, in order to verify the expression changes of the genes upon salt treatment. Expression profiles were compared between the salt tolerant and a susceptible cultivar (Syrah) under both control conditions and after salt stress treatment. Seven cDNA clones were identified which were up-regulated by salt stress in two independent growth experiments and confirmed by RNA blot analysis. The transcript expression patterns of the selected genes differed between the contrasting grapevine cultivars tested with respect to stress-regulation. The possible relationship of individual cDNAs with salinity tolerance mechanisms is discussed.

Genetic diversity in African cucumber (Cucumis sativus L.) provides potential for germplasm enhancement

Genetic diversity among 26 cucumber (Cucumis sativus L. var. sativus) accessions from five African countries [Algeria (1), Egypt (21), Ethiopia (2), Kenya (1), and Libya (1)] present in the U.S. National Plant Germplasm System (NPGS) were examined by assessing variation at 71 polymorphic random amplified polymorphic DNA (RAPD) loci. Genetic distances (GD; simple matching coefficient) were estimated among these African accessions and a reference array (RA) of 21 accessions representative of the genetic variation in cucumber. GD among African accessions ranged between 0.41 and 0.97. GD among accessions in the reference array ranged between 0.36 and 0.88. Multivariate analysis identified three distinct groupings (1–3) of African accessions; Group 1 contained 21 accessions (Egypt, Ethiopia and Libya), Group 2 consisted of two accessions (Kenya, Algeria), and Group 3 possessed three accessions (Egypt). These groupings were distinct from each other (P > 0.001). Accessions in Group 1 differed genetically from all other accessions examined (P > 0.01), and accessions in Groups 2 and 3 were uniquely associated with several RA accessions. While GD among accessions in Group 1 ranged between 0.52 and 0.90, distances among Group 2 accessions varied between 0.93 and 0.97. The GD between the two accessions in Group 3 was 0.65. An accession from Syria (PI 181874) and from one Turkey (PI 199383) were genetically more similar to accessions in Group 1 than to other accessions in the RA. Likewise, accessions in Group 2 were genetically similar to two RA accessions from China and a European glasshouse cucumber line, and Group 3 accessions showed genetic affinities with the U.S. market class cultivar Dasher II. Data suggest that some Egyptian accessions (Group 1) possess unique genetic variation, that this germplasm has potential for broadening the genetic base of commerical cucumber, and that further collection of African germplasm is likely to enhance genetic diversity of cucumber in NPGS.

Investigations on the leaf anatomy and ultrastructure of grapevine (Vitis vinifera) under heat stress

Leaf anatomical and ultrastructural responses of “Razegui” and “Muscat Italia” grapevine cultivars to high temperatures were studied under controlled conditions (T > 36°C), based on photonic and electron microscopy. Histological studies performed on leaves from heat‐stressed and control grapevines revealed thicker leaf blades under high temperature conditions. Environmental scanning electron microscopy of leaf surfaces from both cultivars allowed observing sinuate epidermal cells on the leaves of grapevines cultivated under heat stress and irregular giant oblong pores on their adaxial surface. When observed by transmission electron microscopy, leaf cross sections in grapevines cultivated under high temperature conditions exhibited folded cuticle and cell wall on the adaxial epidermis layer. Therefore, significantly greater cell wall thicknesses were measured under heat stress than control conditions in both cultivars. Regarding chloroplasts, they were more globular in shape under heat stress compared with control conditions and had disorganized thylakoids with a reduced thickness of grana stacking. The size of starch granule decreased, while the number of plastoglobules increased with heat stress, indicating a reduced carbon metabolism and a beginning of senescence within the 3‐month heat stress period. This study confirms widespread adaptive properties in two grapevine cultivars in response to high temperature stress. Microsc. Res. Tech., 2011.

Genetic structure and differentiation among grapevines (Vitis vinifera) accessions from Maghreb region.

Three gene pools representative of Vitis vinifera L. subsp. vinifera (=subsp. sativa Beger) growing in the Maghreb regions (North Africa) from Tunisia (44), Algeria (31) and Morocco (18) and 16 wild grape accessions (Vitis vinifera L. subsp. sylvestris (Gmelin) Beger) from Tunisia were analysed for genetic diversity and differentiation at twenty nuclear microsatellites markers distributed throughout the 19 grape chromosomes. 203 alleles with a mean number of 10.15 alleles per locus were observed in a total of 109 accessions. Genetic diversities were high in all populations with values ranging from 0.6775 (Moroccan cultivars) to 0.7254 (Tunisian cultivars). Fst pairwise values between cultivated grapevine populations were low but found to be significantly different from zero. High Fst pairwise values were shown between wild and cultivated compartments. Two parent offspring relationships, two synonyms and two clones of the same cultivar were detected. The rate of gene flow caused by vegetative dissemination of cultivated grapevine plants was not sufficient to genetically homogenise the pools of cultivars grown in different regions. The Neighbour Joining cluster analysis showed a clear separation according to geographical origins for the cultivated grapevines gene pools and revealed a high dissimilarity between cultivated and wild grapevine. However, three cultivars (Plant d’Ouchtata 1, Plant de Tabarka 3 and Plant d’Ouchtata 3) are very close to wild accessions and may result from a hybridisation between cultivated and wild accessions. The high level of differentiation between cultivated and wild accessions indicates that the cultivated accessions do not derive directly from local wild populations but could mostly correspond to imported materials introduced from others regions during historical times or derived from crossing between them.