Most rice protection methods have currently used toxic chemicals to control pathogens and pests, which leads to environmental pollution. Systemic acquired resistance (SAR) taking advantage of natural defence reaction of plants could be proposed as an alternative, ecologically friendly approach for plant protection. Its application into rice production could minimize the chemicals quantity used and could contribute to the decrease of environmental pollution and the development of sustainable agriculture. The research was conducted to select the most effective chemical and suitable method to improve the health of rice plants infected by grassy stunt disease in net-house of Can Tho University. SAR chemicals were used at very low concentrations (in mM). Results showed that the height of rice plants treated with SAR chemicals was higher than that of plants untreated. Besides, the number of diseased plants was reduced and the ratio of firm grain and yield increased when plants were applied by SAR. Among the used substances, oxalic acid provided the best systemic acquired resistance. With oxalic acid, seed soaking was better than seed coating in systemic acquired resistance against rice grassy stunt disease. / Hầu hết các phương pháp sản xuất lúa hiện nay đều sử dụng các hóa chất độc hại trong việc phòng trừ bệnh và côn trùng gây hại, nên dẫn đến ô nhiễm môi trường. Kích thích tính kháng lưu dẫn giúp kích hoạt cơ chế tự nhiên kháng bệnh của cây có thể là giải pháp bảo vệ thực vật thay thế an toàn với môi trường. Việc ứng dụng tiến bộ này vào trong sản xuất lúa có thể làm giảm lượng hóa chất sử dụng, đóng góp vào việc giảm thiểu ô nhiễm môi trường và sự phát triển của một nền nông nghiệp bền vững. Nghiên cứu đã được thực hiện tại nhà lưới trường Đại học Cần Thơ để tuyển chọn hóa chất và phương pháp sử dụng hóa chất để tăng cường sức khỏe giúp cây lúa vượt qua bệnh vàng lùn. Hóa chất kích kháng được sử dụng ở một nồng độ rất thấp (đơn vị là mM). Kết quả cho thấy chiều cao cây lúa khi xử lý chất kích kháng tốt hơn so đối chứng không xử lý. Bên cạnh đó, số cây lúa nhiễm bệnh giảm, tỉ lệ hạt chắc và năng suất tăng khi cây lúa được xử lý với chất kích kháng. Trong số các chất kích kháng đã sử dụng, acid oxalic cho hiệu quả vượt trội. Với chất acid oxalic, phương pháp ngâm hạt cho hiệu quả kích kháng tốt hơn phương pháp áo hạt.

Systemic acquired resistance (SAR) is initiated upon recognition of specific microbial effectors by cognate plant resistance proteins and immunizes distal tissues of plants against secondary infections. SAR involves the generation of a mobile signal at the site of primary infection, which then translocates to and activates defense responses in the distal tissues via some unknown mechanism(s). This study shows that an ACYL CARRIER PROTEIN 4 (ACP4), GLABRA1 (GL1) and ACYL CARRIER BINDING PROTEINS (ACBP) are required for the processing of the mobile SAR signal in distal tissues of Arabidopsis. Although acp4, gl1 and acbp plants generate the mobile signal, they are unable to respond to this signal to induce systemic immunity. A defective SAR in acp4, gl1 and acbp plants is not associated with salicylic acid (SA)-, methyl SA-, or jasmonic acid-mediated pathways but is related to the presence of an abnormal cuticle on acp4, gl1 and acbp plants. Other genetic mutations impairing the cuticle also compromised SAR. An intact cuticle was only necessary during the time when the mobile signal is generated and translocated to the distal tissues. A novel role for the plant cuticle as the site for SAR-related molecular signaling is demonstrated.

Plants use several strategies to defend themselves against microbial pathogens. These include basal resistance, which is induced in response to pathogen encoded effector proteins, and resistance (R) protein-mediated resistance that is activated upon direct or indirect recognition of pathogen encoded avirulence protein(s). The activation of Rmediated signaling is often associated with generation of a signal, which, upon its translocation to the distal uninfected parts, confers broad-spectrum immunity against related or unrelated pathogens. This phenomenon known as systemic acquired resistance (SAR) is one of the well-established forms of induced defense response. However, the molecular mechanism underlying SAR remains largely unknown. Induction of plant defense is often associated with a fitness cost, likely because it involves reprogramming of the energy-providing metabolic pathways. Glycerol metabolism is one such pathway that feeds into primary metabolism, including lipid biosynthesis. In this study, I evaluated the role of glycerol-3-phosphate (G3P) in host-pathogen interaction. Inoculation with the hemibiotrophic fungal pathogen Colletotrichum higginsianum led to increased accumulation of G3P in wild-type plants. Mutants impaired in biosynthesis of G3P showed enhanced susceptibility, suggesting a correlation between G3P levels and basal defense. Conversely, increased biosynthesis of G3P correlated with enhanced resistance. The Arabidopsis genome encodes one copy of glycerol kinase (GK), which catalyzes phosphorylation of glycerol to G3P, and five copies of G3P dehydrogenase (G3Pdh), which catalyze reduction of dihydroxyacetone phosphate to G3P. Analysis of plants mutated in various G3Pdh's showed that plastidal lipid biosynthesis was only dependent on the GLY1 isoform but the pathogen induced G3P pool required the function of GLY1 and two other G3Pdh isoforms. Interestingly, compromised G3P biosynthesis in GK and G3Pdh mutants also compromised SAR, which was restored when G3P was provided exogenously. Detailed biochemical analysis showed that G3P was transported to distal tissues and that this process was dependent on a lipid transfer protein, DIR1. Together, these results show that G3P plays an important role in both basal- and induced-defense responses.

Nos anos recentes, a atividade agrícola da citricultura vem enfrentando grandes problemas fitossanitários, principalmente, com relação à viabilidade econômica decorrente do controle das doenças. A bactéria Candidatus Liberibacter spp. está associada ao HLB, a principal doença que limita a produção das plantas cítricas. Assim, muitos pesquisadores têm voltado suas atenções para estudarem e encontrarem genes-alvo na resposta do hospedeiro a este patógeno para utilização no melhoramento genético. Nesse sentido, métodos de transformação genética das plantas cítricas são essenciais, porém características inerentes à espécie limitam seu cultivo in vitro e requerem um maior tempo para crescimento e propagação. Com isso, torna-se importante o estudo em plantas modelo, principalmente, para seguir protocolos de validação de genes. De acordo com o exposto, o gene EDS5 isolado de Citrus sinensis, associado ao mecanismo de Resistência Sistêmica Adquirida (SAR) foi superexpresso por meio da transformação genética em tomateiro (Solanum lycopersicum L. Micro-Tom). Após o crescimento dos brotos regenerados, foram identificadas as plantas positivas por meio de análise de GUS e PCR. Linhagens transgênicas homozigotas foram obtidas com avaliação da resistência ao antibiótico canamicina. / In the recent years, the agricultural activity of the citrus industry has been facing big phytosanitary problems, mainly with regard to economic viability arising from disease control. The bacterium Candidatus Liberibacter spp. is associated with HLB, the main disease that limits the production of citrus trees. Thus, many researchers have been returning their attentions to study and find target genes in the host response to this pathogen for use in the genetic improvement. In this way, methods of genetic transformation of citrus plants are essential, but the inherent characteristics of the species border your in vitro cultivation and require a longer time for growth and propagation. Therefore, it is important to study of model plants, mainly for genetic validation protocols. Thus, the EDS5 gene isolated from Citrus sinensis, associated with Systemic Acquired Resistance mechanism (SAR) was overexpressed by genetic transformation in tomato (Solanum lycopersicum L. Micro-Tom). After the growth of regenerated shoots, positive plants were identified by PCR and GUS analysis. Homozygous transgenic lines were obtained with evaluation of resistance to kanamycin.

A cultura da laranja doce é muito importante ao redor do mundo, em especial no Brasil, maior produtor mundial dessas frutas. A produção citrícola sempre esteve ameaçada por muitas doenças de grande importância, tais como, o cancro cítrico, a clorose variegada dos citros (CVC) e pinta preta. Entretanto, em 2004, surgiu o huanglongbing (HLB) ou greening, que tem devastado pomares, e para a qual ainda não foi encontrada uma solução definitiva. A transgenia pode ser uma técnica auxiliar no manejo desta doença com a busca de cultivares mais tolerantes, em especial ao HLB. Neste trabalho, as pesquisas de transgenia não envolveram genes exógenos à planta como, por exemplo, genes de outros organismos ou genes sintéticos, ou seja, foi baseado em tecnologias mais recentes já aplicadas em outras espécies vegetais, nas quais a transgenia é utilizada para super-expressar genes dos sistemas de defesa da própria planta. Estudos indicam que a super-expressão de genes do sistema de Resistência Sistêmica Adquirida (SAR - do inglês, \"Systemic Acquired Resistance\") promove a resistência de plantas a doenças. Um gene importante para esse sistema é o gene npr1 que controla a expressão das proteínas relacionadas à patogênese (PR), em especial a PR1. Junto do gene npr1, os genes npr3 e npr4 também são reguladores desse sistema, atuando sobre o gene npr1 de acordo com os níveis de ácido salicílico presentes na célula, nível este que varia de acordo com o nível de infecção de cada célula. Porém, a avaliação de um evento transgênico de citros pode levar muitos anos. Desta forma, para diminuir esse tempo de avaliação, pensou-se em usar plantas modelos. O sistema escolhido foi o tomateiro Micro-Tom (Solanun lycopersicum L. cv. Micro-Tom). Para a obtenção das construções gênicas, foram identificados os genes Csnpr1, Csnpr3 e Csnpr4 de Citrus sinensis L. Osbeck a partir dos genes Atnpr1, Atnpr3 e Atnpr4 de Arabidopsis thaliana L.. Os genes de citros foram obtidos a partir de uma planta de laranja doce por RT-PCR e clonados no vetor pCambia 2201, que foi então inserido em Agrobacterium tumefaciens para a transformação genética. Foi feita a transformação genética de plantas de laranja doce (Citrus sinensis L. Osbeck) e do tomateiro Micro-Tom. Após o crescimento dos brotos regenerados, foi feita a avaliação das plantas obtidas por meio de PCR. As plantas geneticamente modificadas foram aclimatizadas. As plantas de citros foram enxertadas e mantidas em casa de vegetação. As plantas de tomateiro Micro-Tom foram propagadas por sementes. A progênie foi avaliada aplicando o antibiótico de seleção canamicina, obtendo-se assim uma linhagem transgênica homozigota. / The sweet orange industry is very important worldwide, specially in Brazil, considered the world´s largest producer. The citrus production has always been threatened by several diseases of great importance, such as canker, CVC, and black spot. However, in 2004, the huanglongbing (HLB) or greening has been detected and devastated many citrus groves, and no definitive solution has been found yet. Transgenes may be a helpful tool for the management of this diseases, leading to the production of tolerant cultivars, especially to HLB. In this work, research on transgenic did not include the use of exogenous genes to the plant, such as genes from other organism or synthetic genes, i.e, it was based on new emerging technologies, already used on other crops, in which transgeny is used to super express genes from the plants own defense system. Studies indicate that a super expression of genes from the system called Systemic Acquired Resistance (SAR) promotes disease resistance. One important gene to this system is the npr1 gene, which controls the expression of the pathogen related proteins (PR), in special the PR1. Together with the npr1 gene, the genes npr3 and npr4 are also regulators of this system, regulating the action of the npr1 gene according to the levels of salicylic acid present in the cell, this level varies with the level of infection in each cell. Nevertheless, evaluating a citrus transgenic event may take several years. In order to shorten this time, model plants were used. The model chosen was the Micro-Tom tomato (Solanun lycopersicum L. cv. Micro-Tom). In order to obtain the genetic constructions, the genes Csnpr1, Csnpr3 e Csnpr4 were identified in Citrus sinensis L. Osbeck from the genes, Atnpr1, Atnpr3 and Atnpr4 present in the Arabidopsis thaliana L. genome. The citrus genes were obtained from the citrus genome using RT-PCR procedure and cloned separately into the pCambia 2201 vector, which was inserted into Agrobacterium tumefaciens in order to perform the genetic transformation. Sweet orange (Citrus sinensis L. Osbeck) and Micro-Tom plants were genetically modified. After the growth of the regenerated shoots, the evaluation of the obtained plants was done through PCR analysis. The genetically modified plants were acclimatized, the citrus plants were grafted and kept in the greenhouse, the Micro-Tom plants were propagated trough seeds and its progeny was evaluated by applying the selection antibiotic kanamycin, thus obtaining a homozygous transgenic line.

No description available.

Salicylic acid-binding protein 2 (SABP2) is essential for the establishment of systemic acquired resistance (SAR) in tobacco; SABP2's methyl salicylate (MeSA) esterase activity is required in healthy systemic tissues of infected plants to release the active defense phytohormone SA from MeSA, which serves as a long-distance signal for SAR. In the current study, we characterize a new gene family from Arabidopsis thaliana encoding 18 potentially active α/β fold hydrolases that share 32-57% identity with SABP2. Of 14 recombinant AtMES (MES for methyl esterase) proteins tested, five showed preference for MeSA as a substrate and displayed SA inhibition of MeSA esterase activity in vitro (AtMES1, -2, -4, -7, and -9). The two genes encoding MeSA esterases with the greatest activity, AtMES1 and -9, as well as AtMES7 were transcriptionally upregulated during infection of Arabidopsis with avirulent Pseudomonas syringae. In addition, conditional expression of AtMES1, -7, or -9 complemented SAR deficiency in SABP2-silenced tobacco, suggesting that these three members of the AtMES family are SABP2 functional homologs (orthologs). Underexpression by knockout mutation and/or RNAi-mediated silencing of multiple AtMES genes, including AtMES1, -2, -7, and -9, compromised SAR in Arabidopsis and correlated with enhanced accumulation of MeSA in the systemic tissue of SAR-induced plants. Together, the data show that several members of the AtMES gene family are functionally homologous to SABP2 and redundant for MeSA hydrolysis and probably SAR. These data suggest that MeSA is a conserved SAR signal in Arabidopsis and tobacco.

Systemic acquired resistance (SAR) is a state of heightened defense which is induced throughout a plant by an initial infection; it provides long-lasting, broad-spectrum resistance to subsequent pathogen challenge. Recendy we identified a phloem-mobile signal for SAR which has been elusive for almost 30 years. It is methyl salicylate (MeSA), an inactive derivative of the defense hormone, salicylic acid (SA). This discovery resulted from extensive characterization of SA-binding protein 2 (SABP2), a protein whose high affinity for SA and extremely low abundance suggested that it might be the SA receptor. Instead we discovered that SABP2 is a MeSA esterase whose function is to convert biologically inactive MeSA in the systemic tissue to active SA. The accumulated SA then activates or primes defenses leading to SAR. SABP2's esterase activity is inhibited in the initially/primary infected tissue by SA binding in its active site; this facilitates accumulation of MeSA, which is then translocated through the phloem to systemic tissue for perception and processing by SABP2 to SA. Thus, while SABP2 is not the SA receptor, it can be considered the receptor for the SAR signal. This study of SABPs not only illustrates the unexpected nature of scientific discoveries, but also underscores the need to use biochemical approaches in addition to genetics to address complex biological processes, such as disease resistance.

Salicylic acid is a plant hormone that accumulates with plant-pathogen interaction. This accumulation corresponds to the plant being resistant to infection and without it the plant is susceptible. In this study, primers of genes involved in the normal synthesis of SA were used in RT-PCR to compare gene expression levels in susceptible and resistant plants challenged with tobacco mosaic virus. Because SA synthesis shares chorismate as a common substrate with the synthesis of aromatic amino acids, HPLC was used to determine whether the increase in SA could be attributed to a decrease in amino acid levels. The results suggest that genes of the shikimate pathway are up-regulated in both plant lines but much more quickly in the resistant plant, making differential gene expression a possible cause of SA accumulation. Additionally, results showed a more pronounced decrease in amino acid levels in resistant plants compared to susceptible plants.

<p>Systemic Acquired Resistance (SAR) is a plant defense response induced by an initial infection in one part of the plant that leads to broad-spectrum resistance to normally virulent pathogens in distant naïve leaves. As part of the Cameron research team, I contributed to demonstrating that the lipid transfer protein, DIR1 is required for SAR long distance signaling in <em>Arabidopsis</em> and travels from induced to distant tissues during SAR. A highly similar<em> Arabidopsis</em> protein DIR1-like was identified and is thought to be responsible for the occasional SAR-competent phenotype observed in the <em>dir1-1</em> mutant. This work provides evidence for the idea that DIR1 and DIR1-like are paralogs created by a recent duplication event and that similar to DIR1, DIR1-like may travel to distant tissues during SAR. To better understand DIR1 and DIR1-like contribution during SAR, <em>dir1-1dir1-like</em> double mutant transgenic plants were created as well as transgenic plants expressing epitope- (HA and FLAG) and fluorescent- (iLOV and phiLOV) tagged DIR1 and DIR1-like to facilitate visualization of movement during SAR. Several putative DIR1 orthologs were identified in crop plants and cucumber CucDIR1 was shown to be functionally equivalent to AtDIR1 in <em>dir1-1</em> complementation studies providing further evidence that DIR1 plays an important role in SAR across plant species. By analyzing conservation between DIR1, DIR1-like and the putative DIR1 orthologs, several protein residues were identified that may be important for DIR1 function during SAR. DIR1 proteins were modified at these sites and the importance of these residues was supported by the reduced binding of the TNS hydrophobic probe in these DIR1 variants. Taken together, this thesis suggests that DIR1 and DIR1-like both participate in SAR in <em>Arabidopsis</em>, that DIR1 crop orthologs are also important for the SAR response and that DIR1 possesses several sites that are critical for its function in long distance SAR signaling.</p> / Doctor of Philosophy (PhD)