Identification and Expression Profiles of Six Transcripts Encoding Carboxylesterase Protein in Vitis flexuosa Infected with Pathogens.

Plants protect themselves from pathogen attacks via several mechanisms, including hypersensitive cell death. Recognition of pathogen attack by the plant resistance gene triggers expression of carboxylesterase genes associated with hypersensitive response. We identified six transcripts of carboxylesterase genes, Vitis flexuosa carboxylesterase 5585 (VfCXE5585), VfCXE12827, VfCXE13132, VfCXE17159, VfCXE18231, and VfCXE47674, which showed different expression patterns upon transcriptome analysis of V. flexuosa inoculated with Elsinoe ampelina. The lengths of genes ranged from 1,098 to 1,629 bp, and their encoded proteins consisted of 309 to 335 amino acids. The predicted amino acid sequences showed hydrolase like domains in all six transcripts and contained two conserved motifs, GXSXG of serine hydrolase characteristics and HGGGF related to the carboxylesterase family. The deduced amino acid sequence also contained a potential catalytic triad consisted of serine, aspartic acid and histidine. Of the six transcripts, VfCXE12827 showed upregulated expression against E. ampelina at all time points. Three genes (VfCXE5585, VfCXE12827, and VfCXE13132) showed upregulation, while others (VfCXE17159, VfCXE18231, and VfCXE47674) were down regulated in grapevines infected with Botrytis cinerea. All transcripts showed upregulated expression against Rhizobium vitis at early and later time points except VfCXE12827, and were downregulated for up to 48 hours post inoculation (hpi) after upregulation at 1 hpi in response to R. vitis infection. All tested genes showed high and differential expression in response to pathogens, indicating that they all may play a role in defense pathways during pathogen infection in grapevines.

Preparation of resveratrol-enriched grape juice from ultrasonication treated grape fruits.

Grape (Vitis spp.) is a major source of resveratrol that can be eaten directly or after making jam, jelly, wine and juice. Resveratrol (3,5,4'-trihydroxystilbene) has a profound positive influence on human health, including anti-carcinogenic, anti-cancer, anti-inflammatory, and anti-ageing effects and the ability to lower blood sugar. During industrial production of grape juice, resveratrol is lost because of the use of clarifying agents and filtration; therefore, commercial grape juice contains very low amounts of resveratrol. In this study, we investigated the accumulation of resveratrol in grape juice prepared from three varieties of grape, viz. Campbell Early, Muscat Bailey A (MBA) and Kyoho, following post-harvest ultrasonication cleaning for 5 min and 6h of incubation in the dark at 25 °C. This process resulted in the amounts of resveratrol increasing by 1.53, 1.15 and 1.24 times in juice prepared from Campbell Early, MBA and Kyoho, respectively, without changing the amounts of total soluble solids. Overall, our results indicate that ultrasonication treatment of post-harvested grape fruits can be an effective method for producing resveratrol-enriched grape juice as well as cleaning grapes thoroughly.

Theobroxide Treatment Inhibits Wild Fire Disease Occurrence in Nicotiana benthamiana by the Overexpression of Defense-related Genes.

Theobroxide, a novel compound isolated from a fungus Lasiodiplodia theobromae, stimulates potato tuber formation and induces flowering of morning glory by initiating the jasmonic acid synthesis pathway. To elucidate the effect of theobroxide on pathogen resistance in plants, Nicotiana benthamiana plants treated with theobroxide were immediately infiltrated with Pseudomonas syringae pv. tabaci. Exogenous application of theobroxide inhibited development of lesion symptoms, and growth of the bacterial cells was significantly retarded. Semi-quantitative RT-PCRs using the primers of 18 defense-related genes were performed to investigate the molecular mechanisms of resistance. Among the genes, the theobroxide treatment increased the expression of pathogenesis-related protein 1a (PR1a), pathogenesis-related protein 1b (PR1b), glutathione S-transferase (GST), allen oxide cyclase (AOC), and lipoxyganase (LOX). All these data strongly indicate that theobroxide treatment inhibits disease development by faster induction of defense responses, which can be possible by the induction of defense-related genes including PR1a, PR1b, and GST triggered by the elevated jasmonic acid.

Resistance to Elsinoë ampelina and expression of related resistant genes in Vitis rotundifolia Michx. grapes.

Muscadine grapes (Vitis rotundifolia Michx) are considered as excellent genetic resources for grape breeding programs as they are known for their hardiness and resistance to pests and diseases. However, contrary to popular belief, our study indicated that not all muscadine cultivars are resistant to anthracnose disease. In order to identify a source of genetic tolerance towards anthracnose among muscadine cultivars, a series of in-situ and ex-situ experiments were conducted through strict and sensitive screening processes. Two consecutive years of field evaluation of 54 grape cultivars showed various levels of anthracnose incidence among the cultivars between a scale of 0 (tolerant) to 5 (highly-susceptible). Resistance bioassay by inoculation of different spore densities of Elsinoë ampelina on 40 cultivars presented similar results and was consistent with those obtained from the field test. A real-time PCR analysis was conducted to investigate differences of gene expression between susceptible and tolerant cultivars and to confirm results by phenotypic identification. Expression of genes encoding chalcone synthase, stilbene synthase, polygalacturonase-inhibiting protein, chitinase and lipid transfer-protein was only detected in tolerant cultivars. Resistant muscadine cultivars identified in this study could be excellent candidates for grape disease resistance breeding programs.

Transcriptional profiling of ESTs responsive to Rhizobium vitis from 'Tamnara' grapevines (Vitis sp.).

Genes related with defense responses were screened from the cDNA library constructed with Rhizobium vitis-inoculated or salicylic acid (SA)-treated 'Tamnara' grapevine (Vitis sp.) leaves. Among 13,728 expressed sequence tags (ESTs) from 'Tamnara' grapevine upon R. vitis inoculation and SA treatment, 6776 unigenes containing 1915 contigs and 4860 singletons were obtained. In gene ontology analysis, there were about 3200 clones related with biological process, 3555 with molecular function, and 3354 with cellular component genes. Proteins of secretory organ (35%), plasma membrane (30%), endoplasmic reticulum (20%), and vacuole (11%) were predicted. Photosynthesis-related genes and defense-related genes were most abundant. Among ESTs, 199 resistance-related ones were mapped to the genome of Vitis vinifera L. with three markers, GLP1-12, MHD98, and MHD145, which are known to be linked to resistance against powdery mildew. Approximately, 120 simple sequence repeats (SSRs) detected in cDNAs could be used as EST-derived SSR markers in disease resistant grape breeding.