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Real time qPCR expression analysis of some stress related genes in leaf tissue of Pyrus communis cv. Conférence after infection with Erwinia amylovora
No full textHydrogen peroxide plays a critical role in the
expression of disease resistance in several plant/pathogen
interactions. It serves as a substrate for oxidative crosslinking
of various plant cell wall components leading to the
reinforcement of the cell structure, as a direct toxin against
the pathogen and as a signal molecule for the induction of
defence-related genes in the adjacent, still healthy tissues.
In plant cells, enzymes and redox metabolites act in synergy
to carry out the detoxification of hydrogen peroxide
and other reactive oxygen species (ROS). Superoxide dismutase
(SOD) catalyses the dismutation of superoxide to
hydrogen peroxide, catalase (CAT) dismutates hydrogen
peroxide to oxygen and water, and ascorbate peroxidase
(APX) reduces hydrogen peroxide to water by utilising
ascorbate as specific electron donor. These are considered
some of the main enzymatic systems for protecting cells
against oxidative damage. These enzymes are present in
various isozyme forms in several cell compartments and
their expression is genetically controlled and regulated
both by developmental and environmental stimuli,
according to the necessity to remove ROS produced in
cells. The aim of this study was to determine the possible
role of these antioxidants in the defence mechanism of
Pyrus communis cv. Confe´rence leaf tissue after an infection
with Erwinia amylovora. Shoots of 2-year-old pear
trees cv. Confe´rence were infected with E. amylovora
strain SGB 225/12, were mock infected or left untreated.
To account for structural changes, not only a difference
was made between control, infected and mock-infected
leaves, but we also included a distinction between young
and old leaves, because it is known that older leaves are
less susceptible for fire blight infections. Leaf samples
were taken at specific time points after infection and the
expression pattern of not necrotic tissue close to the
infection site was analysed for their diverse isoforms of
SOD, APX and CAT by using real time qPCR. In this
study, no striking differences in transcription patterns
of these enzymes between control, mock infected and
E. amylovora infected leaves were observed. However, a
significant difference between the expression levels of
some genes in young and old leaves was noticed. These
differences could partially explain the different progression
rate by which E. amylovora infects, invades and causes
necrosis in young and old leaves
Methodological Approach for the Valuation of Biodiversity. Case study: valuation of natural predators for biological pest control in pear production, Flanders
No full textEconomic valuation of natural predators in Pear production
No full textIn spite of global actions, biodiversity is declining at an alarming rate. Despite the need for objectively comparable monetary standards to include biodiversity arguments in policymaking, research on the relationship between species diversity and its valuation from a societal perspective is still scarce.
In this paper, a methodological framework for the valuation of natural predators based on their ecological role in the agroecosystem is introduced. The framework integrates a dynamic ecological model simulating interactions between species with an economic model, thereby quantifying the effect of reduced numbers of natural predators on the net farm income. The model attributes an objective monetary value to increased species diversity through the changes in the provisioning of a marketable good.
Results indicate that the loss of three predators could decrease net farm income with 88.86 €ha-1 to 2186.5 €ha-1. For the pear production sector in Flanders in 2011, this constitutes to an indirect use value of 0,68 million € for one predator and 16.63 million € for the presence of three predators. The aim is to provide a justification for the argument for biodiversity conservation, based on the ecological function of species, through the delivery of comparable monetary standards.The research was made possible with the financial aid from a BOF grant of the Centre for Environmental Sciences (CMK, Hasselt University). Nele Witters is funded by Research Foundation- Flanders (FWO). The authors would like to thank Ellen Elias from Symbio for providing relevant data and insights into the complex interplay between pest insects, natural predators and human impacts from fertilizers and pesticide use
The in planta proteome of wild type strains of the fire blight pathogen, Erwinia amylovora
No full textErwinia amylovora is a Gram-negative plant pathogen that causes fire blight. This disease affects most members of the Rosaceae family including apple and pear. Here, an infection model is introduced to study proteomic changes in a highly virulent E. amylovora strain upon interaction with its host as compared to a lower virulent strain. For this purpose separate shoots of apple rootstocks were wound-infected and when infection became systemic, bacterial cells were isolated and processed for analysis in a proteomics platform combining 2-D fluorescence difference gel electrophoresis and mass spectrometry. Comparing the proteome of the isolates, significant abundance changes were observed in proteins involved in sorbitol metabolism, amylovoran production as well as in protection against plant defense mechanisms. Furthermore several proteins associated with virulence were more abundant in the higher virulent strain. Changes at the proteome level showed good accordance at the transcript level, as was verified by RT-qPCR. In conclusion, this infection model may be a valuable tool to unravel the complexity of plant-pathogen interactions and to gain insight in the molecular mechanisms associated with virulence of E. amylovora, paving the way for the development of plant-protective interventions against this detrimental disease. Significance: During this research a first time investigation was performed on the proteome of E. amylovora, grown inside a susceptible host plant. This bacterium is the causal agent of fire blight, which can affect most members of the Rosaceae family including apple and pear. To do so, an artificial infection model on shoots of apple rootstocks was optimized and employed. When infection was systemic, bacterial cells were extracted from the plant tissue followed by extraction of the proteins from the bacteria. Further processing of the proteins was done by using a 2-D fluorescence difference gel electrophoresis analysis followed by mass spectrometry. By the use of two strains differing in their virulent ability, we were able to draw conclusions concerning virulence and behavior of different strains inside the host. This research provides a model to investigate plant-pathogen interactions and more importantly, we identified possible new targets for the development of novel control methods against this devastating disease. (C) 2016 Elsevier B.V. All rights reserved.Partial funding by project no 101513 of the Agency of Innovation by Science and Technology (IWT-Flanders, Belgium) is acknowledged. Michelle Holtappels was indebted to the IWT for a predoctoral fellowship. We thank Erik Royackers for technical assistance. We acknowledge the financial support from the Hercules Foundation in the framework of the project R-3986 'LC-MS@UHasselt: Linear Trap Quadrupool-Orbitrap mass spectrometer'. We also acknowledge Greet Clerx, Elke Knoops and Annelies Eyskens for their technical support. Dr. Sacha Bohler is acknowledged for his help with the proteomics procedures. The authors have no conflict of interest
Erwinia amylovora affects the phenylpropanoid-flavonoid pathway in mature leaves of Pyrus communis cv. Conference
No full textFlavonoids, which are synthesized by the phenylpropanoid-flavonoid pathway, not only contribute to fruit colour and photoprotection, they also may provide antimicrobial and structural components during interaction with micro-organisms. A possible response of this pathway was assessed in both mature and immature leaves of shoots of 2-year-old pear trees cv. Conference, which were inoculated with the gram-negative bacterium Erwinia amylovora strain SGB 225/12, were mock-inoculated or were left untreated. The phenylpropanoid flavonoid pathway was analysed by histological studies, by gene expression using RT-qPCR and by HPLC analyses of the metabolites at different time intervals after infection. Transcription patterns of two key genes anthocyanidin reductase (ANR) and chalcone synthase (CHS) related to the phenylpropanoid flavonoid pathway showed differences between control, mock-inoculated and E. amy/ovora-inoculated mature leaves, with the strongest reaction 48 h after inoculation. The impact of E. amylovora was also visualised in histological sections, and confirmed by HPLC, as epicatechin which is produced via ANR augmented 72 h after inoculation in infected leaf tissue. Besides the effect of treatments, ontogenesis-related differences were found as well. The increase of certain key genes, the rise in epicatechin and the visualisation in several histological sections in this study suggest a non-negligible impact on the phenylpropanoid flavonoid pathway in Pyrus communis due to inoculation with E. amylovora. In this study, we propose a potential role of this pathway in defence mechanisms, providing a detailed analysis of the response of this system attributable to inoculation with E. amylovora. (C) 2013 Elsevier Masson SAS. All rights reserved.This work was conducted within the framework of the European research network COST864. Partial funding by project no 60686 of the Agency for Innovation by Science and Technology (IWT-Flanders, Belgium) is acknowledged. Kristof Vrancken and Michelle Holtappels are indebted to the IWT for predoctoral fellowships. We are grateful to Jan Daenen, Greet Clerx and Natascha Steffanie for their technical support
Methodological Approach for the Valuation of Biodiversity. Case study: valuation of natural predators for biological pest control in pear production, Flanders
No full textThe plant pathogen Erwinia amylovora: a proteome investigation
No full textErwinia amylovora is considered to be a homogeneous species although different strains show differences in virulence. In this investigation, two wild type strains with differences in virulence, a high virulent strain PFB5 and a low virulent strain LMG2024, were compared using a proteomic approach. Hereby, proteins were
identified which were important for this difference in virulence between both strains. Bacteria were grown in a minimal medium and at mid-exponential phase, samples were taken. The complete proteome was extracted and a 2D differential in-gel electrophoresis (DIGE) approach was used. Differently abundant spots were excised,
trypsinized and identified. Eventually 31 spots were identified as being important in the difference between the two strains. Flagellin (FliC) and a chemotaxis regulatory protein (CheY) were more abundant in the low virulent strain. A protein important in
amylovoran synthesis, UTP-glucose-1-phosphate uridylyltransferase (GalF), was upregulated in the high virulent strain. Afterwards, phenotypic experiments were performed to validate the proteomic results.Partial funding by project no 101513 of the Agency of Innovation by Science and Technology (IWT‐Flanders, Belgium) is acknowledged. Michelle Holtappels is indebted to the IWT for a predoctoral fellowship. We thank Erik Royackers for technical assistance. We acknowledge the financial support from the Hercules Foundation in the framework of the project R‐3986 ‘LC‐MS@UHasselt: Linear Trap Quadrupool‐Orbitrap mass spectrometer’
Economic valuation of natural predators in Pear production
No full textIn spite of global actions, biodiversity is declining at an alarming rate. Despite the need for objectively comparable monetary standards to include biodiversity arguments in policymaking, research on the relationship between species diversity and its valuation from a societal perspective is still scarce.
In this paper, a methodological framework for the valuation of natural predators based on their ecological role in the agroecosystem is introduced. The framework integrates a dynamic ecological model simulating interactions between species with an economic model, thereby quantifying the effect of reduced numbers of natural predators on the net farm income. The model attributes an objective monetary value to increased species diversity through the changes in the provisioning of a marketable good.
Results indicate that the loss of three predators could decrease net farm income with 88.86 €ha-1 to 2186.5 €ha-1. For the pear production sector in Flanders in 2011, this constitutes to an indirect use value of 0,68 million € for one predator and 16.63 million € for the presence of three predators. The aim is to provide a justification for the argument for biodiversity conservation, based on the ecological function of species, through the delivery of comparable monetary standards.The research was made possible with the financial aid from a BOF grant of the Centre for Environmental Sciences (CMK, Hasselt University). Nele Witters is funded by Research Foundation- Flanders (FWO). The authors would like to thank Ellen Elias from Symbio for providing relevant data and insights into the complex interplay between pest insects, natural predators and human impacts from fertilizers and pesticide use
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