Development of a rapid and highly sensitive direct-PCR assay to detect a single conidium of Botrytis cinerea Pers.:Fr in vitro and quiescent forms in planta

“Direct-PCR” amplifications of Botrytis cinerea-specific genomic sequences, without any DNA purification step or time consuming sample preparation, were developed. A single copy sequence of 0.7 Kb in the Botrytis cinerea genome was amplified in reactions containing no more than 1 x 105 to 1 single conidium. As a demonstrative application, this assay was applied to detect B. cinerea in different parts of immature grape berries (at ‘pea size’), when previously inoculated with conidia at flowering. Using this method we showed the presence of quiescent Botrytis in the receptacle area only. Cloning and sequencing of the fragment confirmed the single sequence gene of B. cinerea. These results demonstrate that the method is easy to apply and of sufficiently high sensitivity to detect the presence of B. cinerea in immature grape berries. Its use for studies on the development of grey mould and improved control of the disease in vineyards is discussed

Effects of resveratrol, viniferins and pterostilbene on Plasmopara viticola zoospore mobility and disease development

The effects of stilbenes (resveratrol, δ- and ε-viniferins, and pterostilbene) on the mobility of zoospores of Plasmopara viticola and on subsequent disease development were studied in vitro. δ-viniferin and pterostilbene are the most toxic stilbenes concerning zoospore mobility (ED50 : 14.6 and 28.3 μM) and disease development (ED50 : 14.7 and 12.7 μM). The analysis of stilbenes in leaf cells of resistant (Solaris) and susceptible (Chasselas) grape cultivars artificially inoculated with P. viticola has shown that very high amounts of stilbenic phytoalexins accumulate at the site of infection of the resistant cultivar compared to the susceptible one

Histological and biochemical criteria for objective and early selection of grapevine cultivars resistant to Plasmopara viticola

Grapevine breeding is the most effective way to create cultivars resistant to downy mildew (Plasmopara viticola), and to reduce the number of fungicide applications. Four criteria, including histological and biochemical analyses, based on the level of different mechanisms of resistance to grapevine downy mildew, were tested on 42 different cultivars. Plantlets were artificially inoculated with downy mildew and the sporangia density was measured spectrophotometrically 6 d after infection. Callose synthesis in stomata and δ- and ε-viniferin levels at the site of infection were recorded 48 h after inoculation. These observations have allowed the 42 cultivars to be divided into 5 groups: very resistant (VR), resistant (R), less susceptible (LS), susceptible (S) and highly susceptible (HS). All 4 criteria have to be applied to assign the resistance level closer to field conditions. This method allows to rapidly evaluate the level of resistance of seedlings to downy mildew thereby leading to a reduction in duration  of the breeding program by several years.

Compared efficacy of preservation solutions in liver transplantation: A long-term graft outcome study from the european liver transplant registry

International audienceBetween 2003 and 2012, 42 869 first liver transplantations performed in Europe with the use of either University of Wisconsin solution (UW; N = 24 562), histidine-tryptophan-ketoglutarate(HTK; N = 8696), Celsior solution (CE; N = 7756) or Institute Georges Lopez preservation solution (IGL-1; N = 1855) preserved grafts. Alternative solutions to the UW were increasingly used during the last decade. Overall, 3-year graft survival was higher with UW, IGL-1 and CE (75%, 75% and 73%, respectively), compared to the HTK (69%) (p 12 h or grafts used for patients with cancer (p < 0.0001). For partial grafts, 3-year graft survival was 89% for IGL-1, 67% for UW, 68% for CE and 64% for HTK (p = 0.009). Multivariate analysis identified HTK as an independent factor of graft loss, with recipient HIV (+), donor age ≥65 years, recipient HCV (+), main disease acute hepatic failure, use of a partial liver graft, recipient age ≥60 years, no identical ABO compatibility, recipient hepatitis B surface antigen (-), TIT ≥ 12 h, male recipient and main disease other than cirrhosis. HTK appears to be an independent risk factor of graft loss. Both UW and IGL-1, and CE to a lesser extent, provides similar results for full size grafts. For partial deceased donor liver grafts, IGL-1 tends to offer the best graft outcome

Evaluation of Hungarian Wines for Resveratrol by Overpressured Layer Chromatography

A method, including solid phase extraction sample preparation, overpressured layer chromatographic separation and subsequent densitometric evaluation, was developed for measurement of total resveratrol (cis- and trans-isomers) content of wine. The amount of resveratrol was determined in wine samples from different winemaking regions of Hungary. The total resveratrol was high in Hungarian red wines (3.6–11 mg/L), and much lower in white ones (0.04–1.5 mg/L)

Transient Receptor Potential Channel Polymorphisms Are Associated with the Somatosensory Function in Neuropathic Pain Patients

Transient receptor potential channels are important mediators of thermal and mechanical stimuli and play an important role in neuropathic pain. The contribution of hereditary variants in the genes of transient receptor potential channels to neuropathic pain is unknown. We investigated the frequency of transient receptor potential ankyrin 1, transient receptor potential melastin 8 and transient receptor potential vanilloid 1 single nucleotide polymorphisms and their impact on somatosensory abnormalities in neuropathic pain patients. Within the German Research Network on Neuropathic Pain (Deutscher Forscbungsverbund Neuropathischer Schmerz) 371 neuropathic pain patients were phenotypically characterized using standardized quantitative sensory testing. Pyrosequencing was employed to determine a total of eleven single nucleotide polymorphisms in transient receptor potential channel genes of the neuropathic pain patients and a cohort of 253 German healthy volunteers. Associations of quantitative sensory testing parameters and single nucleotide polymorphisms between and within groups and subgroups, based on sensory phenotypes, were analyzed. Single nucleotide polymorphisms frequencies did not differ between both the cohorts. However, in neuropathic pain patients transient receptor potential ankyrin 1 710G>A (rs920829, E179K) was associated with the presence of paradoxical heat sensation (p = 0.03), and transient receptor potential vanilloid 1 1911A>G (rs8065080, I585V) with cold hypoalgesia (p = 0.0035). Two main subgroups characterized by preserved (1) and impaired (2) sensory function were identified. In subgroup 1 transient receptor potential vanilloid 1 1911A>G led to significantly less heat hyperalgesia, pinprick hyperalgesia and mechanical hypaesthesia (p = 0.006, p = 0.005 and p<0.001) and transient receptor potential vanilloid 1 1103C>G (rs222747, M315I) to cold hypaesthesia (p = 0.002), but there was absence of associations in subgroup 2. In this study we found no evidence that genetic variants of transient receptor potential channels are involved in the expression of neuropathic pain, but transient receptor potential channel polymorphisms contributed significantly to the somatosensory abnormalities of neuropathic pain patients

Metabolic constituents of grapevine and grape-derived products

The numerous uses of the grapevine fruit, especially for wine and beverages, have made it one of the most important plants worldwide. The phytochemistry of grapevine is rich in a wide range of compounds. Many of them are renowned for their numerous medicinal uses. The production of grapevine metabolites is highly conditioned by many factors like environment or pathogen attack. Some grapevine phytoalexins have gained a great deal of attention due to their antimicrobial activities, being also involved in the induction of resistance in grapevine against those pathogens. Meanwhile grapevine biotechnology is still evolving, thanks to the technological advance of modern science, and biotechnologists are making huge efforts to produce grapevine cultivars of desired characteristics. In this paper, important metabolites from grapevine and grape derived products like wine will be reviewed with their health promoting effects and their role against certain stress factors in grapevine physiology

Effects of Clinically Relevant MPL Mutations in the Transmembrane Domain Revealed at the Atomic Level through Computational Modeling

BACKGROUND: Mutations in the thrombopoietin receptor (MPL) may activate relevant pathways and lead to chronic myeloproliferative neoplasms (MPNs). The mechanisms of MPL activation remain elusive because of a lack of experimental structures. Modern computational biology techniques were utilized to explore the mechanisms of MPL protein activation due to various mutations. RESULTS: Transmembrane (TM) domain predictions, homology modeling, ab initio protein structure prediction, and molecular dynamics (MD) simulations were used to build structural dynamic models of wild-type and four clinically observed mutants of MPL. The simulation results suggest that S505 and W515 are important in keeping the TM domain in its correct position within the membrane. Mutations at either of these two positions cause movement of the TM domain, altering the conformation of the nearby intracellular domain in unexpected ways, and may cause the unwanted constitutive activation of MPL's kinase partner, JAK2. CONCLUSIONS: Our findings represent the first full-scale molecular dynamics simulations of the wild-type and clinically observed mutants of the MPL protein, a critical element of the MPL-JAK2-STAT signaling pathway. In contrast to usual explanations for the activation mechanism that are based on the relative translational movement between rigid domains of MPL, our results suggest that mutations within the TM region could result in conformational changes including tilt and rotation (azimuthal) angles along the membrane axis. Such changes may significantly alter the conformation of the adjacent and intrinsically flexible intracellular domain. Hence, caution should be exercised when interpreting experimental evidence based on rigid models of cytokine receptors or similar systems