Project description:Clubroot of Brassicaceae, an economically important soil borne disease, is caused by Plasmodiophora brassicae Woronin, an obligate, biotrophic protist. This disease poses a serious threat to canola and related crops in Canada and around the globe causing significant loss to seed yield. The pathogen is continuously evolving and new pathotypes are emerging, this necessitates the development of novel resistant canola cultivars to manage the disease effectively. Given that proteins play a crucial role in majority of biological processes and molecular functions, the identification of differentially abundant proteins (DAP) using proteomics information is an attractive approach to understand the plant-pathogen interactions as well as in the future development of gene specific markers for developing clubroot resistant (CR) cultivars. In this study, P. brassicae pathotype 3 (P3H) was used to challenge CR and clubroot susceptible (CS) canola lines. Root samples were collected at three distinct stages of pathogenesis, 7-, 14-, and 21-days post inoculation (DPI), protein samples were isolated, digested with trypsin and subjected to LC-MS/MS analysis. A total of 937 proteins demonstrated a significant (q < 0.05) change in abundance in at least in one of the time points when compared between control and inoculated CR-parent, CR-progeny, CS-parent, CS-progeny and 784 proteins were significantly (q < 0.05) changed in abundance in at least in one of the time points when compared between the inoculated- CR and CS root proteomes of parent and progeny across the three time points tested. Functional annotation of the differentially abundant proteins (DAPs) revealed several proteins related to calcium dependent signaling pathways in response to the pathogen. In addition, proteins related to reactive oxygen species (ROS) biochemistry, dehydrins, lignin, thaumatin, and phytohormones were identified. Among the DAPs, 74 putative proteins orthologous to CR proteins and quantitative trait loci (QTL) associated with eight CR loci in four chromosomes including chromosomes A3 and A8 were identified. In conclusion, these results have contributed to an improved understanding of the mechanisms that are involved in mediating response to P. brassicae in canola at the protein level.
Project description:The hemibiotrophic fungal pathogen Leptosphaeria maculans is the causal agent of blackleg disease in Brassica napus (canola, oilseed rape) and causes significant losses in crop yields worldwide. While genetic resistance has been used to mitigate the disease, little information about the genes and gene regulatory networks underlying blackleg resistance is currently available. High-throughput RNA sequencing and rigorous bioinformatics approaches revealed dynamic changes in the host transcriptome and identified plant defense pathways specific to the host-pathogen incompatible LepR1-AvrLepR1 interaction.
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 5’ 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.
Project description:Background: The biological control agent Pseudomonas chlororaphis PA23 is effective at protecting Brassica napus (canola) from the necrotrophic fungus Sclerotinia sclerotiorum via direct antagonism. Despite the growing importance of biocontrol bacteria in plant protection from fungal pathogens, little is known about how the host plant responds to bacterial priming on the leaf surface or about changes in gene activity genome-wide in the presence and absence of S. sclerotiorum. Results: PA23 priming of mature canola plants reduced the number of lesion forming petals by 90%. Global RNA sequencing of the host pathogen interface showed a reduction in the number of genes uniquely upregulated in response to S. sclerotiorum by 16-fold when pretreated with PA23. Upstream defense-related gene patterns suggest MAMP-triggered immunity via surface receptors detecting PA23 flagellin and peptidoglycans. Although systemic acquired resistance was induced in all treatment groups, a response centered around a glycerol-3-phosphate (G3P)-mediated pathway was exclusively observed in plants treated with PA23 alone. Activation of these defense mechanisms by PA23 involved mild reactive oxygen species production as well as pronounced thylakoid membrane structures and plastoglobule formation in leaf chloroplasts. Conclusion: Further to the direct antibiosis that it exhibits towards the pathogen S. sclerotiorum, PA23 primes defense responses in the plant through the induction of unique local and systemic defense regulatory networks. This study has shed light on the potential effects of biocontrol agents applied to the plant phyllosphere. Understanding these interactions will aid in the development of biocontrol systems as a viable alternative to chemical pesticides in the protection of important crop systems. Mature canola leaf tissue treated with combinations of PA23 or S. sclerotiorum ascospores (3 treatment groups) was compared to a water treated control (all treatments done in triplicate).
Project description:Cultivation of canola at temperatures above the optimum growth temperature of 21°C for prolonged periods, especially during the flowering stage, resulted in several adverse effects, including rapid vegetative growth, reduced viability of female gametophytes, increased seed abortion rate, accelerated embryo development, and a reduction in seed oil composition (Young et al., 2004; Mácová et al., 2022; Secchi et al., 2023). One of the distinctive phenotypes observed during the seed development of certain canola cultivars subjected to prolonged heat stress affecting the seed yield is the occurrence of the pre-harvest sprouting phenotype (PHS) (Mácová et al., 2022). Misregulation of seed dormancy by abscisic acid and dormancy-related genes is thought to be the primary cause of PHS in many cereal crops (Benech-Arnold & Rodríguez, 2018; Tai et al., 2021). This phenotype is associated with seed coat rupture (SCR), observed in seeds during the early stages of maturation. In this study, we employed a multi-methodological approach to investigate the occurrence of SCR phenotype in seeds of Brassica napus cv. Topas. The results demonstrate that SCR occurs in seeds around 20 days after pollination (20DAP) when the plants are cultivated in elevated temperatures over an extended period. The unrestricted embryonic growth exerts pressure on the seed coat, as evidenced by a reduction in the thickness of the seed coat cell layers. This results in an early alteration to the cell wall composition, with an increased proportion of demethylesterified pectin, which is likely to stiffen the seed coat, thereby rendering it more susceptible to rupture. The precise mechanism by which accelerated embryo development influences heat stress-mediated seed development in canola plants has yet to be elucidated.
Project description:Background: The biological control agent Pseudomonas chlororaphis PA23 is effective at protecting Brassica napus (canola) from the necrotrophic fungus Sclerotinia sclerotiorum via direct antagonism. Despite the growing importance of biocontrol bacteria in plant protection from fungal pathogens, little is known about how the host plant responds to bacterial priming on the leaf surface or about changes in gene activity genome-wide in the presence and absence of S. sclerotiorum. Results: PA23 priming of mature canola plants reduced the number of lesion forming petals by 90%. Global RNA sequencing of the host pathogen interface showed a reduction in the number of genes uniquely upregulated in response to S. sclerotiorum by 16-fold when pretreated with PA23. Upstream defense-related gene patterns suggest MAMP-triggered immunity via surface receptors detecting PA23 flagellin and peptidoglycans. Although systemic acquired resistance was induced in all treatment groups, a response centered around a glycerol-3-phosphate (G3P)-mediated pathway was exclusively observed in plants treated with PA23 alone. Activation of these defense mechanisms by PA23 involved mild reactive oxygen species production as well as pronounced thylakoid membrane structures and plastoglobule formation in leaf chloroplasts. Conclusion: Further to the direct antibiosis that it exhibits towards the pathogen S. sclerotiorum, PA23 primes defense responses in the plant through the induction of unique local and systemic defense regulatory networks. This study has shed light on the potential effects of biocontrol agents applied to the plant phyllosphere. Understanding these interactions will aid in the development of biocontrol systems as a viable alternative to chemical pesticides in the protection of important crop systems.