Brassica nigra challenged by Pieris brassicae, eggs of Pieris brassicae, Xanthomonas campestris pv. raphani, Brevicoryne brassicae or by combined stresses eggs of P. brassicae / P. brassicae, X. campestris / P. brassicae, B. brassicae / P. brassicae
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ABSTRACT: Brassica nigra plants, a Brassicaceae close to Arabidopsis thaliana, was used for combined stresses experiments. In this study, we performed a whole-genome microarray analysis on five-week-old plants and compared untreated plants and plants treated different single or dual stresses: the larvae Pieris brassicae, egg extract of Pieris brassicae, the bacterial Xanthomonas campestris pv. raphani, the aphid Brevicoryne brassicae or by combined stresses eggs of P. brassicae / P. brassicae, X. campestris / P. brassicae, B. brassicae / P. brassicae.
Project description:Brassica nigra plants, a Brassicaceae close to Arabidopsis thaliana, was used for combined stresses experiments. In this study, we performed a whole-genome microarray analysis on five-week-old plants and compared untreated plants and plants treated with ozone at 70 ppb, larvae of Pieris brassicae or both ozone followed by P. brassicae insect.
Project description:Plants can cope with stress better if they experience a mild form of the stress before the actual \\"real\\" stress event. In Arabidopsis thaliana it is known that plants that harboured eggs of the White cabbage butterfly (Pieris brassicae) before larval feeding can defend better against the herbivore stress. The main aim of the experiment was to compare the priming effect induced by insect egg deposition of Pieris brassicae between vegetative and reproductive (first open flowers) Arabidopsis thaliana plants on the transcriptional level. We used a full factorial setup consisting of a) untreated control plants , b) plants which experienced eggs for 6 days without larval feeding after that period, c) plants which experienced no eggs before larval feeding for 24 hours d) plants which experienced eggs for 6 days and larval herbivory for 24 hours. This setup was conducted with 6 week old vegetative plants and 10 week old reproductive plants were the first flowers were open. For all treatments leaf tissue from the leaves that experienced egg oviposition and/or larval feeding were collected. From reproductive plants flower buds were collected as well.
Project description:Plant defence against insects is well known to be affected by previous exposure to cues warning of herbivory. Using Arabidopsis thaliana and the herbivore Pieris brassicae, we addressed the question whether the maintenance of the effects of the warning cue depends on its reliability. We determined the transcriptomes of Arabidopsis leaves that were treated by P. brassicae egg deposition (i) five days after oviposition, (ii) one day after removal of the eggs following the egg treatment, (iii) three days after removal of the eggs, (iv) after two days of herbivory that started one day after removal of the eggs; or that were treated by chilling (v) five days after transfer to 4°C, (vi) one day after transfering the plants to 20°C following the chilling treatment, (vii) three days after transfering the plants to 20°C, (viii) after two days of herbivory that started one day after transfering the plants to 20°C. Arabidopsis thaliana Col-0 wild type plants were grown under short day conditions (10 h/14 h light/dark) at 20°C for 7 weeks. Subsequently (i) the plants were transferred to 4°C for 5 days, or (ii) Pieris brassicae deposited ca. 40 eggs on leaf 17 where they remained for five days, or (iii) as controls plants grew untreated for five days. Next, the plants were transferred back to 20°C and the eggs were removed, respectively. Next, all plants rested for 1 day at 20°C. Next, P. brassicae larvae were allowed to feed for 2 days on leaf 17 adjacent to the former egg deposit site or at a respective leaf region of chilling-treated or untreated plants. Control plants were not exposed to larvae. From all treated and untreated plants material from a leaf region proximal to the egg deposition and/or feeding site was harvested for transcriptome analysis.
Project description:In Arabidopsis thaliana it is known that plants that harboured eggs of the White cabbage butterfly (Pieris brassicae) before larval feeding can defend better against the herbivore stress. As Arabidopsis thaliana is an annual plant we were interested in the plant defense and possible priming reaction of the closest biannual/perennial relative Arabidopsis lyrata. The main aim of the experiment was to compare the effect induced by insect egg deposition of Pieris brassicae between A. thaliana and A. lyrata plants grown in parallel on the transcriptional level. We used a full factorial setup consisting of a) untreated control plants , b) plants which experienced eggs for 6 days without larval feeding after that period, c) plants which experienced no eggs before larval feeding for 24 hours d) plants which experienced eggs for 6 days and larval herbivory for 24 hours. This setup was conducted with 7 week old vegetative plants For all treatments leaf tissue from the leaves that experienced egg oviposition and/or larval feeding were collected.
Project description:Arabipdosis thaliana (ecotype Col-0) was infected with the root pathogen Plasmodiophora brassicae. Gene expression of the host plant has been analyzed at two time points after inoculation (10 and 23 days after inoculation) compared to the correspondend control plants.
Project description:Canola (oilseed rape, Brassica napus L.), is susceptible to infection by the biotrophic protist Plasmodiophora brassicae, the causal agent of clubroot. To understand the roles of microRNAs (miRNAs) during the post-transcriptional regulation of disease initiation and progression, we have characterized the changes in miRNA expression profiles in canola roots during clubroot disease development and have compared these to uninfected roots. Two different stages of clubroot development were targeted in this miRNA profiling study: an early time of 10-dpi for disease initiation and a later 20-dpi, by which time the pathogen had colonized the roots (as evident by visible gall formation and histological observations). P. brassicae responsive miRNAs were identified and validated by qRT-PCR of miRNAs and the subsequent validation of the target mRNAs through starBase degradome analysis, and through 5M-bM-^@M-^Y RLM-RACE. This study identifies putative miRNA-regulated genes with roles during clubroot disease initiation and development. Putative target genes identified in this study included: transcription factors (TFs), hormone-related genes, as well as genes associated with plant stress response regulation such as cytokinin, auxin/ethylene response elements. The results of our study may assist in elucidating the role of miRNAs in post-transcriptional regulation of target genes during disease development and may contribute to the development of strategies to engineer durable resistance to this important phytopathogen. In this miRNA-microarray experiment a total of 4 samples were analyzed with their 3 biological replicates. In which 2 samples C 10 DAY and C 20 DAY was used as referrence contols.
Project description:Clubroot (Plasmodiophora brassicae) is a pathogen of Brassicaceae that causes significant reductions in yield as a consequence of gall formation in the root and hypocotyl of infected plants. The pathogen hijacks host vascular cambium development and cytokinins are implicated in this process. RNASeq was used to investigate changes in cytokinin metabolism during gall formation of clubroot-infected Arabidopsis thaliana. RNASeq analysis of infected tissue showed that host cytokinin metabolism was strongly down-regulated both at the onset (16 DPI) and late (26 DPI) stages of gall formation. Expression of host genes associated with cytokinin biosynthesis, signalling, degradation and conjugation was strongly repressed. Two isopentenyltransferase genes associated with cytokinin biosynthesis are present in the P. brassicae genome and are expressed throughout gall formation.
Project description:The clubroot disease caused by the obligate biotrophic protist Plasmodiophora brassicae on host plants of the Brassicaceae family is characterized by enhanced cell division and cell expansion. Since a typical root section of an infected plant always includes different stages of the pathogen as well as uninfected cells, we were interested to investigate specific developmental stages of the pathogen and their effect on host transcriptional changes. We extended previous microarray studies on whole roots by using Laser Microdissection and Pressure Catapulting (LMPC) to isolate individual cells harboring defined developmental stages of the pathogen. In addition, we compared the central cylinder of infected to contol plants. We were especially interested to elucidate the stage-specific hormonal network. The upregulation of genes involved in auxin and cytokinin metabolism and signaling was confirmed. In addition, we found evidence that brassinosteroid (BR) synthesis and signal perception was in many cases upregulated in enlarged cells and the central cylinder. This was confirmed by qPCR and mutant analysis of the BR receptor mutant bri1-6, which exhibited less severe gall formation than the respective wild type. Our results identify novel hormone pathways involved in clubroot development. Using this method of single cell preparation combined with transcriptome analysis has been very useful to elucidate the regulation of gall growth by this obligate biotropic pathogen in a cell- and stage-specific manner. Transcription profiling was performed in isolated Arabidopsis thaliana root cells harboring different developmental stages of Plasmodiophora brassicae at two time points after inoculation (dai) (14 and 21 dai), as well as in infected central cylinder tissue from roots at 14 dai (days after inoculation). Control samples were taken from uninfected roots. Host cells were dissected from paraffin embedded roots using Laser Microdissection and Pressure Catapulting (LMPC). 8 samples have been analyzed.
Project description:Clubroot (Plasmodiophora brassicae) is a pathogen of Brassicaceae that causes significant reductions in yield as a consequence of gall formation in the root and hypocotyl of infected plants. The pathogen manipulates host cell cycle regulation in order to drive proliferation in early stages of infection and endoreduplication in later stages. Microarrays were used to profile changes in the expression of cell cycle control components at the onset (16 DPI) and late (26 DPI) stages of gall formation.
Project description:We have implemented an integrated Systems Biology approach to analyze overall transcriptomic reprogramming and systems level defense responses in the model plant Arabidopsis thaliana during an insect (Brevicoryne brassicae) and a bacterial (Pseudomonas syringae pv. tomato strain DC3000) attack. The main aim of this study was to identify the attacker-specific and general defense response signatures in the model plant Arabidopsis thaliana while attacked by phloem feeding aphids or pathogenic bacteria. Defense responses and networks, unique and specific for aphid or Pseudomonas stresses were identified. Our analysis revealed a probable link between biotic stress and microRNAs in Arabidopsis and thus opened up a new direction to conduct large-scale targeted experiments to explore detailed regulatory links among them. The presented results provide a first comprehensive understanding of Arabidopsis - B. brassicae and Arabidopsis - P. syringae interactions at a systems biology level. Arabidopsis thaliana (ecotype Colombia-0) seeds were grown in 6-cm-diameter pots filled with a sterile soil mix (1.0 part soil and 0.5 part horticultural perlite), 3 plants per pot. Plants were kept in growth chambers VM-CM-6tsch VB 1514 (VM-CM-6tsch Industrietechnik GmbH, Germany) under the following conditions: a 8/16 h (light/dark) photoperiod at 22M-BM-0C/18M-BM-0C, 40%/70% relative humidity, and 70/0 M-NM-<mol m-2s-1 light intensity. After 32 days plants had 8 fully developed leaves. Each plant was infested with 32 wingless aphids [Brevicoryne Brassicae] (4 per leaf), which were transferred to leaves with a fine paintbrush. Infested plants and aphid-free controls were kept in plexiglass cylinders. Plants were harvested 72 h after infestation between the 6th and 8th hour of the light photoperiod. Four biological replicates were prepared from aphid infested and control plants, each sampled from 15 individual plants. Whole rosettes were cut at the hypocotyls and aphids were removed by washing with Milli-Q-filtered water. Differences in transcriptional responses were measured by comparing genes expression of aphid infested plants against non-infested control plants.