Elucidating the interaction between Brassica napus and Rhizoctonia solani AG 2-1

Brassica napus, oilseed rape (OSR), is a worldwide cultivated crop belonging to the family Brassicaceae, broadly used in crop rotations with cereals. Production is focused on oil for human consumption, biodiesel and feedstock. OSR has undergone intensive breeding for optimization of oil content, disease resistance and augmentation of yields, and today is considered one of the most profitable crops. Nonetheless, oilseed rape is the primary host for the necrotrophic soil-borne pathogen Rhizoctonia solani anastomosis group (AG) 2-1. Infection of seedlings causes damping off disease and decreases crop establishment and yields. AG 2-1 is the most prevalent AG of R. solani in wheat fields in the UK. Currently there is no OSR germplasm resistant to R. solani AG 2-1. Available control methods include cultural practices and chemical seed treatments, which aim to postpone the infection and hence improve crop establishment. Changes in agronomic practices and crop management, including choice of cultivars, tillage, application of fertilisers and pesticides, mean that there is a danger of future outbreaks of diseases that in the past were not considered as major problem. This includes R. solani AG 2-1 which can infect other rotational crops as well and due to its saprophytic nature remains in the fields for years. The aim of the PhD was to elucidate interactions between R. solani AG 2-1 and B. napus, by identifying potential resistant traits and understanding how the pathogen counteracts OSR plant defences. The first objective was to develop and compare different high-throughput screening methods that could be used for the phenotyping of OSR germplasm interactions with R. solani AG 2-1. Four methods were developed and compared: (1) nutrient media plates, (2) compost trays, (3) light expanded clay aggregate (LECA) trays and (4) a hydroponic pouch and wick system. Inoculation of LECA was the most suitable method for screening disease caused by AG 2-1 to OSR germplasm, because it allowed the detection of differences in disease severity between the tested OSR genotypes 5 days post infection (dpi) and also to conduct measurements in whole plants. The second objective was to identify any sources of disease resistance by screening a diversity of OSR germplasm. To start the screening, I selected randomly germplasm from commercial cultivars and parental lines of mapping populations that was available in our seed bank. Overall, the germplasm tested consisted of commercial cultivars, genotypes from diversity sets and a mapping population. All genotypes tested appeared to be susceptible to AG 2-1 infection as shown by high disease levels, reduced emergence and survival. Additionally, I tested if any induced defence responses from exposure to disease could be inherited in the next generation through an epigenetic stress response. However, all progeny plants were also highly susceptible indicating that there was no evidence for transgenerational induction of resistance in this system. The third objective was to gain insight into OSR plant defences when exposed to a combination of attacking organisms, as this often occurs in real field situations. I investigated the role of M. persicae infestation on OSR susceptibility to R. solani AG 2-1. There was no effect of AG 2-1 infection on aphid performance. However, M. persicae infestation resulted in significantly more disease symptoms in B. napus cv. ‘Canard’ plants although there were no significant differences in the amount of fungal DNA. Marker genes LOX3 and MYC2 had an augmented expression under AG 2-1 treatment but were downregulated in plants exposed to both aphids and pathogen. Hence, it appears that aphid infestation induced changes in the jasmonic acid (JA) signalling pathway, which resulted in the increased susceptibility to AG 2-1. In conclusion, the present work provided a new high-throughput screening method suitable to phenotype disease by AG 2-1 in the early seedling stage within a short time period. Unfortunately, the current results confirm previous studies indicating that AG 2-1 is an extremely aggressive isolate to OSR germplasm that lacks genetic resistance. Nonetheless, the observed differences between the germplasm tested in the present work suggest that there are potential tolerant traits. For the first time, the current work provided evidence that M. persicae infestation can negatively affect plant defences against R. solani AG 2-1, through suppression of genes involved in JA signalling. Additionally, it was demonstrated that R. solani AG 2-1 induces the activation of defence mechanism related to both JA and salicylic acid (SA) pathways. Future studies aiming to identify resistant/tolerant traits should screen wider Brassica germplasm, including wild species. Additionally, it will be particularly interesting to explore how R. solani overcomes OSR defences by examining the expression of a broader array of genes involved in plant defence mechanisms

Elucidating the interaction between Brassica napus and Rhizoctonia solani AG 2-1

Brassica napus, oilseed rape (OSR), is a worldwide cultivated crop belonging to the family Brassicaceae, broadly used in crop rotations with cereals. Production is focused on oil for human consumption, biodiesel and feedstock. OSR has undergone intensive breeding for optimization of oil content, disease resistance and augmentation of yields, and today is considered one of the most profitable crops. Nonetheless, oilseed rape is the primary host for the necrotrophic soil-borne pathogen Rhizoctonia solani anastomosis group (AG) 2-1. Infection of seedlings causes damping off disease and decreases crop establishment and yields. AG 2-1 is the most prevalent AG of R. solani in wheat fields in the UK. Currently there is no OSR germplasm resistant to R. solani AG 2-1. Available control methods include cultural practices and chemical seed treatments, which aim to postpone the infection and hence improve crop establishment. Changes in agronomic practices and crop management, including choice of cultivars, tillage, application of fertilisers and pesticides, mean that there is a danger of future outbreaks of diseases that in the past were not considered as major problem. This includes R. solani AG 2-1 which can infect other rotational crops as well and due to its saprophytic nature remains in the fields for years. The aim of the PhD was to elucidate interactions between R. solani AG 2-1 and B. napus, by identifying potential resistant traits and understanding how the pathogen counteracts OSR plant defences. The first objective was to develop and compare different high-throughput screening methods that could be used for the phenotyping of OSR germplasm interactions with R. solani AG 2-1. Four methods were developed and compared: (1) nutrient media plates, (2) compost trays, (3) light expanded clay aggregate (LECA) trays and (4) a hydroponic pouch and wick system. Inoculation of LECA was the most suitable method for screening disease caused by AG 2-1 to OSR germplasm, because it allowed the detection of differences in disease severity between the tested OSR genotypes 5 days post infection (dpi) and also to conduct measurements in whole plants. The second objective was to identify any sources of disease resistance by screening a diversity of OSR germplasm. To start the screening, I selected randomly germplasm from commercial cultivars and parental lines of mapping populations that was available in our seed bank. Overall, the germplasm tested consisted of commercial cultivars, genotypes from diversity sets and a mapping population. All genotypes tested appeared to be susceptible to AG 2-1 infection as shown by high disease levels, reduced emergence and survival. Additionally, I tested if any induced defence responses from exposure to disease could be inherited in the next generation through an epigenetic stress response. However, all progeny plants were also highly susceptible indicating that there was no evidence for transgenerational induction of resistance in this system. The third objective was to gain insight into OSR plant defences when exposed to a combination of attacking organisms, as this often occurs in real field situations. I investigated the role of M. persicae infestation on OSR susceptibility to R. solani AG 2-1. There was no effect of AG 2-1 infection on aphid performance. However, M. persicae infestation resulted in significantly more disease symptoms in B. napus cv. ‘Canard’ plants although there were no significant differences in the amount of fungal DNA. Marker genes LOX3 and MYC2 had an augmented expression under AG 2-1 treatment but were downregulated in plants exposed to both aphids and pathogen. Hence, it appears that aphid infestation induced changes in the jasmonic acid (JA) signalling pathway, which resulted in the increased susceptibility to AG 2-1. In conclusion, the present work provided a new high-throughput screening method suitable to phenotype disease by AG 2-1 in the early seedling stage within a short time period. Unfortunately, the current results confirm previous studies indicating that AG 2-1 is an extremely aggressive isolate to OSR germplasm that lacks genetic resistance. Nonetheless, the observed differences between the germplasm tested in the present work suggest that there are potential tolerant traits. For the first time, the current work provided evidence that M. persicae infestation can negatively affect plant defences against R. solani AG 2-1, through suppression of genes involved in JA signalling. Additionally, it was demonstrated that R. solani AG 2-1 induces the activation of defence mechanism related to both JA and salicylic acid (SA) pathways. Future studies aiming to identify resistant/tolerant traits should screen wider Brassica germplasm, including wild species. Additionally, it will be particularly interesting to explore how R. solani overcomes OSR defences by examining the expression of a broader array of genes involved in plant defence mechanisms

Infestation by Myzus persicae Increases Susceptibility of Brassica napus cv. “Canard” to Rhizoctonia solani AG 2-1

Activation of plant defense pathways can be influenced by the presence of different species of attacking organisms. Understanding the complicated interactions triggering plant defense mechanisms is of great interest as it may allow the development of more effective and sustainable disease control methods. Myzus persicae and Rhizoctonia solani anastomosis group (AG) 2-1 are two important organisms attacking oilseed rape (OSR), causing disease and reduced yields. At present, is unclear how these two interact with each other and with OSR defenses and therefore the aim of the present study was to gain a better insight into the indirect interaction between aphids and pathogen. In separate experiments, we assessed the effect of AG 2-1 infection on aphid performance, measured as growth rate and population increase and then the effect of aphid infestation on AG 2-1 by quantifying disease and the amount of fungal DNA in plant stems and compost for two OSR varieties, “Canard” and “Temple.” Additionally, we examined the expression of genes related to jasmonic acid (JA) and salicylic acid (SA) defense pathways. There was no significant effect of AG 2-1 infection on M. persicae performance. However, aphid infestation in one of the varieties, “Canard,” resulted in significantly increased disease symptoms caused by AG 2-1, although, the amount of fungal DNA was not significantly different between treatments. This meant that “Canard” plants had become more susceptible to the disease. Expression of LOX3 and MYC2 was elevated under AG 2-1 treatment but downregulated in plants with both aphids and pathogen. Therefore it seems plausible that alterations in the JA signaling due to aphid infestation resulted in the increased susceptibility to AG 2-1

Development of high-throughput methods to screen disease caused by Rhizoctonia solani AG 2-1 in oilseed rape

Background: Rhizoctonia solani (Kühn) is a soil-borne, necrotrophic fungus causing damping off, root rot and stem canker in many cultivated plants worldwide. Oilseed rape (OSR, Brassica napus) is the primary host for anastomosis group (AG) 2-1 of R. solani causing pre- and post-emergence damping-off resulting in death of seedlings and impaired crop establishment. Presently, there are no known resistant OSR genotypes and the main methods for disease control are fungicide seed treatments and cultural practices. The identification of sources of resistance for crop breeding is essential for sustainable management of the disease. However, a high-throughput, reliable screening method for resistance traits is required. The aim of this work was to develop a low cost, rapid screening method for disease phenotyping and identification of resistance traits. Results: Four growth systems were developed and tested: (1) nutrient media plates, (2) compost trays, (3) light expanded clay aggregate (LECA) trays, and (4) a hydroponic pouch and wick system. Seedlings were inoculated with virulent AG 2-1 to cause damping-off disease and grown for a period of 4–10 days. Visual disease assessments were carried out or disease was estimated through image analysis using ImageJ. Conclusion: Inoculation of LECA was the most suitable method for phenotyping disease caused by R. solani AG 2-1 as it enabled the detection of differences in disease severity among OSR genotypes within a short time period whilst allowing measurements to be conducted on whole plants. This system is expected to facilitate identification of resistant germplasm

The Loss of α- and β-Tubulin Proteins Are a Pathological Hallmark of Chronic Alcohol Consumption and Natural Brain Ageing

Repetitive excessive alcohol intoxication leads to neuronal damage and brain shrinkage. We examined cytoskeletal protein expression in human post-mortem tissue from Brodmann’s area 9 of the prefrontal cortex (PFC). Brain samples from 44 individuals were divided into equal groups of 11 control, 11 alcoholic, 11 non-alcoholic suicides, and 11 suicide alcoholics matched for age, sex, and post-mortem delay. Tissue from alcoholic cohorts displayed significantly reduced expression of α- and β-tubulins, and increased levels of acetylated α-tubulin. Protein levels of histone deacetylase-6 (HDAC6), and the microtubule-associated proteins MAP-2 and MAP-tau were reduced in alcoholic cohorts, although for MAPs this was not significant. Tubulin gene expressions increased in alcoholic cohorts but not significantly. Brains from rats administered alcohol for 4 weeks also displayed significantly reduced tubulin protein levels and increased α-tubulin acetylation. PFC tissue from control subjects had reduced tubulin protein expression that was most notable from the sixth to the eighth decade of life. Collectively, loss of neuronal tubulin proteins are a hallmark of both chronic alcohol consumption and natural brain ageing. The reduction of cytosolic tubulin proteins could contribute to the brain volumetric losses reported for alcoholic patients and the elderly

Pathologists and entomologists must join forces against forest pest and pathogen invasions

The world’s forests have never been more threatened by invasions of exotic pests and pathogens, whose causes and impacts are reinforced by global change. However, forest entomologists and pathologists have, for too long, worked independently, used different concepts and proposed specific management methods without recognising parallels and synergies between their respective fields. Instead, we advocate increased collaboration between these two scientific communities to improve the long-term health of forests. Our arguments are that the pathways of entry of exotic pests and pathogens are often the same and that insects and fungi often coexist in the same affected trees. Innovative methods for preventing invasions, early detection and identification of non-native species, modelling of their impact and spread and prevention of damage by increasing the resistance of ecosystems can be shared for the management of both pests and diseases. We, therefore, make recommendations to foster this convergence, proposing in particular the development of interdisciplinary research programmes, the development of generic tools or methods for pest and pathogen management and capacity building for the education and training of students, managers, decision-makers and citizens concerned with forest health.The European Union’s Horizon 2020 research and innovation programmehttp://neobiota.pensoft.netam2021BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Pathologists and entomologists must join forces against forest pest and pathogen invasions

The world’s forests have never been more threatened by invasions of exotic pests and pathogens, whose causes and impacts are reinforced by global change. However, forest entomologists and pathologists have, for too long, worked independently, used different concepts and proposed specific management methods without recognising parallels and synergies between their respective fields. Instead, we advocate increased collaboration between these two scientific communities to improve the long-term health of forests. Our arguments are that the pathways of entry of exotic pests and pathogens are often the same and that insects and fungi often coexist in the same affected trees. Innovative methods for preventing invasions, early detection and identification of non-native species, modelling of their impact and spread and prevention of damage by increasing the resistance of ecosystems can be shared for the management of both pests and diseases. We, therefore, make recommendations to foster this convergence, proposing in particular the development of interdisciplinary research programmes, the development of generic tools or methods for pest and pathogen management and capacity building for the education and training of students, managers, decision-makers and citizens concerned with forest health