Project description:Application of the 'Boerner' (Vitis riparia X Vitis cinerea) genome assembly for targeted mapping of the downy mildew resistance Rpv14 in Vitis.
Project description:Plasmopara viticola (Berk. and Curt.) Berl. and de Toni is the agent of the destructive disease known as grapevine downy mildew, for the control of which intensive fungicide treatments are required. Natural sources of resistance are available in several wild Vitis species, which are being used in traditional breeding approaches. However, molecular switches, signals and effectors involved in resistance are poorly understood. In this paper we report a microarray analysis of early transcriptional changes associated to P. viticola infection in both susceptible Vitis vinifera and resistant Vitis riparia plants (12 and 24 h post inoculation). To provide a biological basis to the choice of time points for transcriptome analyses, we performed microscopic examinations of infected tissues at 12, 24, 48 and 96 hpi. Data suggest that resistance in V. riparia is mainly a post-infectional event and involves a large reprogramming of host metabolism. Transcripts of signal transduction-related genes are specifically and often strongly accumulated in response to infection. Well known defence genes also show marked transcript increases, especially pathogenesis-related proteins PR-10 and stylbene synthases, and genes related to an hypersensitive reaction. On the other hand, V. vinifera mounts a much weaker transcriptional response, involving mainly defence genes, not effective enough in preventing pathogen infection.
Project description:Plasmopara viticola (Berk. and Curt.) Berl. and de Toni is the agent of the destructive disease known as grapevine downy mildew, for the control of which intensive fungicide treatments are required. Natural sources of resistance are available in several wild Vitis species, which are being used in traditional breeding approaches. However, molecular switches, signals and effectors involved in resistance are poorly understood. In this paper we report a microarray analysis of early transcriptional changes associated to P. viticola infection in both susceptible Vitis vinifera and resistant Vitis riparia plants (12 and 24 h post inoculation). To provide a biological basis to the choice of time points for transcriptome analyses, we performed microscopic examinations of infected tissues at 12, 24, 48 and 96 hpi. Data suggest that resistance in V. riparia is mainly a post-infectional event and involves a large reprogramming of host metabolism. Transcripts of signal transduction-related genes are specifically and often strongly accumulated in response to infection. Well known defence genes also show marked transcript increases, especially pathogenesis-related proteins PR-10 and stylbene synthases, and genes related to an hypersensitive reaction. On the other hand, V. vinifera mounts a much weaker transcriptional response, involving mainly defence genes, not effective enough in preventing pathogen infection. Leaves from one resistant (V. riparia cv. Gloire de Montpellier) and one susceptible (V.vinifera cv. Pinot Noir) grapevine cultivars grown in vitro were infected with the oomycete Plasmopara viticola, and transcriptome changes were investigated at 12h and 24h after infection. Three biological replicates were considered and each hybridization was performed twice. One color labeling was performed
Project description:Study of gene expression during Plasmopara viticola infection in the resistant Vitis vinifera cultivar 'Regent'. The oomycete fungus Plasmopara viticola (Berk. et Curt.) Berl. et de Toni is responsible for grapevine downy mildew disease. Most of the cultivated grapevines are sensitive to this pathogen, thus requiring intensive fungicide treatments. The molecular basis of resistance to this pathogen is poorly understood. We have carried out a cDNA microarray transcriptome analysis to identify grapevine genes associated with resistance traits. Early transcriptional changes associated with downy mildew infection in the resistant Vitis vinifera cultivar ‘Regent’, when compared to the susceptible cultivar ‘Trincadeira’, were analyzed. Transcript levels were measured at three time-points: 0, 6 and 12 hours post inoculation (hpi). Our data indicate that resistance in V. vinifera ‘Regent’ is induced after infection. This study provides the identification of several candidate genes that may be related to ‘Regent’ defense mechanisms, allowing a better understanding of this cultivar's resistance traits.
Project description:Grapevine downy mildew caused by the Oomycete Plasmopara viticola is one of the most important diseases affecting Vitis spp. However, all cultivated European grapevine varieties are susceptible to P. viticola and the resistance needs to be introduced from other Vitaceae. Segregating populations derived from Muscadinia rotundifolia, a species closely related to Vitis, lead to the identification and mapping of two resistance genes, named Rpv1 and Rpv2. The macroscopic phenotypes of the resistance mediated by the two loci are different. Rpv2 plants completely inhibit P. viticola sporulation and produce very small necrotic lesions. In contrast, Rpv1 plants allow a rather limited but visible sporulation of P. viticola. The aim of the study is to understand the gene expression changes associated with downy mildew resistance mediated by these two loci. Gene expression patterns after P. viticola inoculation or mock inoculation are compared in incompatible (resistant plants bearing Rpv1 or Rpv2 locus) and compatible (susceptible plants with no resistance loci) interactions, using the Vitis vinifera GeneChip from Affymetrix. In order to limit the effect of the genetic background, the plant material consists in three pools of genotypes called B, C and D, respectively corresponding to partially resistant plants (Rpv1+/Rpv2-), totally resistant plants (Rpv1-/Rpv2+), and susceptible plants (Rpv1-/Rpv2-) derived from a segregating population. Each pool consists in 3 different genotypes. Plant leaf discs were inoculated with P. viticola sporangium suspension (i) or mock-inoculated with water (ni) and analysed 6 hours post-inoculation. For each experimental condition, we performed two biological replicates. Keywords: normal vs disease comparison
Project description:Grapevine downy mildew caused by the Oomycete Plasmopara viticola is one of the most important diseases affecting Vitis spp. However, all cultivated European grapevine varieties are susceptible to P. viticola and the resistance needs to be introduced from other Vitaceae. Segregating populations derived from Muscadinia rotundifolia, a species closely related to Vitis, lead to the identification and mapping of two resistance genes, named Rpv1 and Rpv2. The macroscopic phenotypes of the resistance mediated by the two loci are different. Rpv2 plants completely inhibit P. viticola sporulation and produce very small necrotic lesions. In contrast, Rpv1 plants allow a rather limited but visible sporulation of P. viticola. The aim of the study is to understand the gene expression changes associated with downy mildew resistance mediated by these two loci. Gene expression patterns after P. viticola inoculation or mock inoculation are compared in incompatible (resistant plants bearing Rpv1 or Rpv2 locus) and compatible (susceptible plants with no resistance loci) interactions, using the Vitis vinifera GeneChip from Affymetrix. In order to limit the effect of the genetic background, the plant material consists in three pools of genotypes called B, C and D, respectively corresponding to partially resistant plants (Rpv1+/Rpv2-), totally resistant plants (Rpv1-/Rpv2+), and susceptible plants (Rpv1-/Rpv2-) derived from a segregating population. Each pool consists in 3 different genotypes. Plant leaf discs were inoculated with P. viticola sporangium suspension (i) or mock-inoculated with water (ni) and analysed 6 hours post-inoculation. For each experimental condition, we performed two biological replicates. Experiment Overall Design: 3 plant genotypes were analyzed: Rpv1+/Rpv2- (B), Rpv1-/Rpv2+ (C), and Rpv1-/Rpv2- (D). Plants were either inoculated with P. viticola (i) or mock-inoculated with water (ni). Biological replicates were performed for each experimental condition.
Project description:Study of gene expression during Plasmopara viticola infection in the resistant Vitis vinifera cultivar 'Regent'. The oomycete fungus Plasmopara viticola (Berk. et Curt.) Berl. et de Toni is responsible for grapevine downy mildew disease. Most of the cultivated grapevines are sensitive to this pathogen, thus requiring intensive fungicide treatments. The molecular basis of resistance to this pathogen is poorly understood. We have carried out a cDNA microarray transcriptome analysis to identify grapevine genes associated with resistance traits. Early transcriptional changes associated with downy mildew infection in the resistant Vitis vinifera cultivar M-bM-^@M-^XRegentM-bM-^@M-^Y, when compared to the susceptible cultivar M-bM-^@M-^XTrincadeiraM-bM-^@M-^Y, were analyzed. Transcript levels were measured at three time-points: 0, 6 and 12 hours post inoculation (hpi). Our data indicate that resistance in V. vinifera M-bM-^@M-^XRegentM-bM-^@M-^Y is induced after infection. This study provides the identification of several candidate genes that may be related to M-bM-^@M-^XRegentM-bM-^@M-^Y defense mechanisms, allowing a better understanding of this cultivar's resistance traits. 3 time points: 0, 6 and 12 hours post inoculation by P. viticola. Two cultivars: control (Trinacedira) and test (Regent). Two biological replicates were performed at 0 hpi, and 3 biological replicates at 6 and 12hpi. At 12hpi, three technical replicates also were performed.
Project description:European grapevine cultivars (Vitis vinifera spp.) are highly susceptible to the downy mildew pathogen Plasmopara viticola. Breeding of resistant V. vinifera cultivars is a promising strategy to reduce the impact of disease management. Most cultivars that have been bred for resistance to downy mildew, rely on resistance mediated by the Rpv3 (Resistance to P. viticola) locus. However, despite the extensive use of this locus, little is known about the mechanism of Rpv3-mediated resistance. In this study, Rpv3-mediated defense responses were studied in Rpv3+ and Rpv3ˉ grapevine cultivars following inoculation with two distinct P. viticola isolates avrRpv3+ and avrRpv3ˉ, with the latter being able to overcome Rpv3 resistance. Based on comparative microscopic, metabolomic and transcriptomic analyses, our results show that the Rpv3-mediated resistance is associated with a defense mechanism that triggers synthesis of fungi-toxic stilbenes and programmed cell death (PCD), resulting in reduced but not suppressed pathogen growth and development. Functional annotation of the encoded protein sequence of genes significantly upregulated during the Rpv3-mediated defense response revealed putative roles in pathogen recognition, signal transduction and defense responses.
Project description:A comparative proteomic analysis of grapevine leaves from the resistant genotype V. davidii ‘LiuBa-8’ (LB) and susceptible genotype V. vinifera ‘Pinot Noir’ (PN) at 12 hpi was conducted to understand the complex relationship between grapevine and P. viticola and the molecular mechanisms between difference resistant vitis genotypes at the early stage of infection. A total of 444 and 349 differentially expressed proteins (DEPs) were identified in LB and PN respectively at 12 hpi by iTRAQ. MapMan analysis showed that the majority of these DEPs were related to photosynthesis, metabolism, stress and redox. More up-expressed DEPs involved in photosynthesis and less down-expressed DEPs involved in metabolism contribute to the resistance in LB. PR10.2, PR10.3, HSF70.2 and HSP90.6 are proposed to be key proteins in both compatible and incompatible interactions. Accumulation H2O2 established the ROS signaling in incompatible interaction and APXs, GSTs maybe associated with the resistance of grapevine against downy mildew. Moreover, we verified four proteins to ensure the accuracy of proteome data using PRM. Overall, these data provide new insights into molecular events and provide valuable candidate proteins that could be used to illuminate molecular mechanisms underlying the incompatible and compatible interaction in early stage and eventually to be exploited to develop new protection strategy against downy mildew in grapevine.
Project description:A comparative proteomic analysis of grapevine leaves from the resistant genotype V. davidii ‘LiuBa-8’ (LB) and susceptible genotype V. vinifera ‘Pinot Noir’ (PN) at 12 hpi was conducted to understand the complex relationship between grapevine and P. viticola and the molecular mechanisms between difference resistant vitis genotypes at the early stage of infection. A total of 444 and 349 differentially expressed proteins (DEPs) were identified in LB and PN respectively at 12 hpi by iTRAQ. MapMan analysis showed that the majority of these DEPs were related to photosynthesis, metabolism, stress and redox. More up-expressed DEPs involved in photosynthesis and less down-expressed DEPs involved in metabolism contribute to the resistance in LB. PR10.2, PR10.3, HSF70.2 and HSP90.6 are proposed to be key proteins in both compatible and incompatible interactions. Accumulation H2O2 established the ROS signaling in incompatible interaction and APXs, GSTs maybe associated with the resistance of grapevine against downy mildew. Moreover, we verified four proteins to ensure the accuracy of proteome data using PRM. Overall, these data provide new insights into molecular events and provide valuable candidate proteins that could be used to illuminate molecular mechanisms underlying the incompatible and compatible interaction in early stage and eventually to be exploited to develop new protection strategy against downy mildew in grapevine.