Systemic acquired resistance (SAR): A novel strategy for plant protection.
- 3. Facts Process of activating natural defense system present in plants induced by biotic or abiotic factors.Kothari, 2004 Sensitize plants to respond rapidly after infection. Kuc and Tuzun, 1990 Regulate genes for defense compounds, effect is systemic and often lasts for the life of annual plants. Kuc, 1990 Increases the levels of : Chitinases β -1,3- glucanases Peroxidases. Ye et al., 1989 Base of immunization : Expression of potential of resistance. Tuzun, 1990
- 4. Systemic Acquired Resistance Systemic acquired resistance (SAR) was first recognized as a significant phenomenon in the early1900s. Upon infection with necrotizing pathogens many plants develop an enhanced resistance to further pathogen attack also in the uninoculated organs. This type of enhanced resistance is referred to as systemic acquired resistance (SAR). Ross (1961) coined this induced resistance in plants as systemic Acquired resistance (SAR) and described it as a long lasting, broad spectrum, Systemic resistance to subsequent infections induced by a pathogen.
- 5. Historical background Van Loon and Van Kammen (1970) and Gianinazzi et al.(1970) showed that pathogenic infection of tobacco induced the accumulation of a distinct set of proteins, called pathogenesis related proteins (PR proteins). Cai et al. (1996) observed that when rice seedlings were sprayed at 0.01 MM of salicylic acid, blast severity was reduced by 59.8% compared with water treated control. Ross (1961) coined this resistance in plants as systemic Acquired resistance (SAR) and described it as a long lasting, broad spectrum, Systemic resistance. The first controlled laboratory study of SAR was performed by Ross (1961) demonstrated that inoculation of a single leaf of tobacco with tobacco mosaic virus (TMV) reduced the severity of subsequent infections on other leaves. White (1979) found that acetyl salicylic acid application is sufficient to induce PR gene expression and enhanced resistance to TMV. Klessig and Raskin led to hypothesis that SA accumulates in cucumber and tobacco plants prior to pathogen infection and found that SA acts as endogenous signal molecule that is required for SAR induction. Truman et al. (2007) have revealed that systemic accumulation of SA during the onset of SAR is preceded by a variety of metabolic signals Such Jasmonates, indole-derived compounds (Truman et al. (2010).
- 6. Salicylic Acid (SA) COOH OH Accumulates in both local and systemic tissues (not the systemic signal) Removal of SA (as in nahG plants) prevents induction of SAR Analogs: INA or BTH
- 7. Systemic Acquired Resistance Systemic acquired resistance (SAR) is an important component of plant defense against pathogen infection. Systemic acquired resistance (SAR) confers long-lasting protection against a broad spectrum of microorganisms. SAR requires the signal molecule salicylic acid (SA) and is associated with accumulation of pathogenesis-related.
- 8. SAR Chemicals Park et al. 2007 Salicylic acid (SA) and its methylated derivative (MeSA) Truman et al. 2007 Jasmonic acid (JA) Truman et al. 2010 Auxin Navarova et al. 2012 Pipecolic acid (Pip) Chaturvedi et al. 2012 Dehdryoabietinal (DA) Jung et al. 2009 Azelaic acid (AzA) Jakab et al. 2001 BABA 2,6-dichloroisonicotinic acid (INA) benzothiadiazole (BTH) inoculation of an avirulent pathogen. Natural elicitors (PGPR).
- 9. Structure of SAR Chemicals
- 10. SA-The endogenous signal in SAR It has been proposed that salicylic acid acts as an endogenous signal responsible for inducing systemic acquired resistance in plants (Gozzo and Faoro, 2013). The contribution of salicylic acid to systemic acquired resistance was investigated in transgenic tobacco plants harboring a bacterial gene encoding salicylate hydroxylase which converts salicylic acid to catechol. Transgenic plants that express salicylate hydroxylase accumulated little or no salicylic acid and were defective in their ability to induce acquired resistance against tobacco mosaic virus. Thus, salicylic acid is essential for the development of systemic acquired resistance.
- 11. Time requirement The onset of SAR involves the generation of mobile signal(s) at the site of local infection. But it may take also days for SAR to develop throughout the host plant (Kuc, 1982). The signal(s) then translocate through the phloem to the systemic tissues where it activates defence response. This occurs within 4–6 hours of primary infection (Chaturvedi et al., 2012).
- 12. Signal Transduction Pathway Specific resistance is based on pathogen-receptor binding Signal transduction pathway is triggered Antimicrobial molecules that seal off infected areas are produced Infected cells release methyl salicylic acid before they die The signaling molecule is distributed to rest of the plant In cells away from infection site, methyl salicylic acid is converted to salicylic acid which initiates a signal transduction pathway Molecules that help protect the cell against a diversity of pathogens for several days are produced
- 14. Salicylic acid (SA) biosynthesis in plants
- 16. AzelaicAcid Pathway Inhibit subsequent infection Antimicrobial defenses Accumulates very high levels of salicylic acid Induces production of AZI1 Azelaic acid increases Primary infection (Sproales, 2012)
- 17. Proteins who have roles in SA synthesis Recently identified PBS3 and EPS1. PBS3 encodes a member of the acyl adenylate/thioester-forming enzyme family. EPS1 encodes a member of the BAHD acyltransferase superfamily. PBS3 and EPS1 are involved in the synthesis of an important precursor or regulatory molecule for SA biosynthesis.
- 18. Involvement of H2O2 in SAR Elevated level of cellular reactive oxygen species (i.e., H2O2) was observed in plants with SAR. Chen and Klessig (1993) isolated a SA-binding protein (SABP) from the cDNA library of tobacco leaves. The deduced cDNA sequence indicates that SABP is a catalase. They proposed that SA activates the elevated levels of cellular H2O2 by inhibiting the catalase activity. Cellular H2O2 functions as a secondary messenger in SAR signal transduction.
- 19. SABP Catalase H2O2 H2O + O2 Catalase-SA SA Disease resistance Defense genes (Klessig, 1993) (SA-binding protein) INA BTH
- 20. Mechanism of SAR Van Loon and Van Kammen(1970) and Gianinazzi et al.(1970) showed that pathogenic infection of tobacco induced the accumulation of a distinct set of proteins, called pathogenesis related proteins(PR proteins).It can be also induced by synthetic resistance inducing compounds. In response to SAR, the positive regulator protein NPR1 moves to the nucleus where it interacts with TGA transcription factors to induce defense gene expression, thus activating SAR (Durrant and Dong 2004). Ward et al demonstrated SAR genes. SAR genes product have direct antimicrobial activity closely related to classes of anti microbial proteins. These includes • Chitinases • Beta 1,3 glucanases • Cysteine rich proteins.
- 21. Hypersensitive response Rapid death of cells in the local region surrounding an infection. Restrict the growth and spread of pathogens to other parts of the plant. Favour growth of pathogens with a necrotrophic lifestyle.
- 22. Phytoalexins The term Phytoalexin is derived from Greek- phyto meaning plant and alexin means warding off compound. Muller (1958) demonstrated the detection of a chemical entity as a Phytoalexin while working with the hypersensitive response of bean tissue Phytoalexins are defined as low molecular weight, anti- microbial compounds that are both synthesized and accumulated in plants after exposure to microorganisms or abiotic agents (Mert- Turk, 2002) They are broad spectrum inhibitors and are chemically diverse with different type’s characteristic of particular plant species. Phytoalexins tend to fall into several classes including terpenoids, glycosteroids and alkaloids. Derivatives simple phenylpropanoid pathway, Shikimic acid pathway, Trp pathway and mevalonic acid pathway (Hammerschmidt, 1999)
- 24. Gene for gene hypothesis For each resistance gene in the host there is a corresponding gene for avirulence in the pathogen cnferring resistance and viceversa
- 25. The gene-for-gene resistance model Host Genes MicrobeGenes RR or Rr rr Avr1 No disease Disease avr1 Disease Disease
- 26. SAR Avr R gene SA NPR1 PR-1 PR-2 PR-5 SAR
- 27. ETI and PTI
- 28. Salicylic acid pathway Production of active oxygen Thickening of plant cell wall PR-proteins, Antimicrobial compounds Production of phenolic esters Salicylic acid(SA)
- 29. Production of pathogenesis related proteins PR protein are plant protein that are induced in pathological and related situation. These proteins are accumulated 7-10 days after infection and indicate the attainment of SAR. It is accumulated in the intercellular spaces(first line of defence) and vacuole (second line of defence by lytic enzyme).
- 32. Structural features of NPR1 593 amino acids, 67 kD Two protein-protein interaction domains: BTB/POZ and Ankyrin repeats Contains NLS Multiple phosphorylation sites No DNA binding domain npr 1-1 BTB ARD S S NLSnpr 1-2 nim 1-2
- 33. PR proteins Plants in which PRP detected Enzymes Target pathogen sites PR 1 Rice, barley, maize, tomato, tobacco Active against glucan activity Plant cell wall thickening Resistance to the spread of the pathogen on the apoplast PR 2 Rice, barley, maize, tomato, tobacco, potato, pepper, bean, Brassica, sugar beet β-1-3-glucanase Cell wall glucan of fungi PR 3 Rice, maize, tomato, pepper, sugar beet, rape seed Chitinase Cell wall chitin of fungi PR 4 Tomato, tobacco, rubber tree Chitinase Cell wall chitin of fungi PR 5 Rice, wheat, barley, oats, tomato, tobacco, potato Thaumatine- like Against oomycetes PR 6 barley, tomato, tobacco Proteinase inhibitor Against nematodes and insects PR 7 Tomato Endoproteinase Microbial cell wall dissolution PR 8 Cucumber Chitinase with lysozyme activity Cell wall chitin of fungi and mucopepetide of bacteria PR 9 Tomato, rice, tobacco, wheat Peroxidase Indirect anti microbial activity as cross linking of plant cell wall PR 10 Potato, asperagus, pea, bean, rice Ribonucleases Viral RNA PR 11 Tobacco Chitinase Cell wall chitin of fungi PR 12 Arabidopsis, pea Defensin Antifungal and antibacterial PR 13 Barley Thionin Antifungal and antibacterial PR 14 Barley Lipid transfer proteins Antifungal and antibacterial PR 15 Barley oxalate oxidase H2O2 production PR 16 Barley and wheat Oxalate-oxidase like with superdismutase activity H2O2 production PR 17 Wheat, barley, tobacco Peptidase (Van Loon, 2006)
- 34. PR protein in mulberry WAP18 extracted from cortical parenchyma cells of mulberry tree. Cryoprotective effects on the freeze-labile enzyme, LDH. WAP18 may function in the freezing tolerance mechanism of cortical parenchyma cells of mulberry tree during winter. (Ukaji et al., 2004)
- 35. Laticifer A laticifer is an elongated tubular cell and its cytoplasmic content (latex) is thought to be involved in defense against herbivores and microbes. Insecticidal chitinase-like proteins (LA-a and b) unlignified tissues Antifungal class I chitinase (LA-c) lignified tissues. Mulberry laticifers accumulate large amounts of biotic-stress-related defense proteins e.g., pathogenesis-related protein-1, β-1-3-glucanase, class V chitinase, osmotin and lectins. They are adapted to different threats. (Kitajima et al., 2013)
- 36. Unlignified tissues Lignified tissues Class V chitinase β-1,3-glucanase β-1,3-glucanase Class I chitinase Class I chitinase Lectin Class I chitinase galactose-binding Osmotin Lectin, galactose-binding Pathogenesis-related protein 1 Lectin, mannose-binding (Kitajima et al., 2013)
- 37. Chitinase genes in mulberry plants Mulberry genome encodes 20 chitinase genes. Expression of mulberry chitinase genes in five different tissues including root, bark, bud, flower, and leaf. Detect transcriptional differences in their response to Insect wounding Fungal infection Biochemical elicitors Found that chitinase genes were induced by insect wounding and fungal infection. Thus play an important role in plant defense responses. Accumulation of Mnchi16 transcripts in the insect- wounded leaves. Mnchi8 and Mnchi19 were upregulated in response to infection. Mnchi19 strongly inhibit the hyphal extension of the fungus. Mnchi8 and Mnchi16 were strongly induced by different factors, suggesting that different mulberry chitinases respond to different biotic stresses. This information will be crucial in advancing our understanding of mulberry chitinase genes for Sericultural improvement (Wang et al., 2015)
- 38. Gene Name Accession No. Mnchi1 Morus022978 Mnchi2 Morus007185 Mnchi3 Morus007186 Mnchi4 Morus017594 Mnchi5 Morus022481 Mnchi6 Morus022482 Mnchi7 Morus020088 Mnchi8 Morus011486 Mnchi9 Morus011484 Mnchi10 Morus003149 Mnchi11 Morus020224 Mnchi12 Morus000037 Mnchi13 Morus007737 Mnchi14 Morus012010 Mnchi15 Morus018118 Mnchi16 Morus018119 Mnchi17 Morus018124 Mnchi18 Morus013887 Mnchi19 Morus014360 Mnchi20 Morus014362
- 39. Defensins Defensins are small cationic peptides of 45-54 amino acid residues with anti-microbial activities. They inhibit the growth of wide range of phytopathogenic microorganisms. They are the new member of thionine family. Defensins are expressed in most but not all plants and are key members of a plants immune system. Defensins provide a first line of defense against pathogen attack. Defensins work at level of innate nonspecific immunity against the varied pathogens. Nontoxic to most animal and plant cells. Defenins are thermostable. Defensins have shown satisfactory efficacy against pathogens
- 40. History 1995 1990 First plant defensin was isolated by Mendez and colleagues. Plant defensin termed was coined by Terras and colleagues 40 1995 Reported the use of defensin to enhance the resistance to fungal pathogen by Terras and colleagues
- 41. Biological activity Biological activity Antibacterial Antifungal α-amylases and serine proteinases Zinc tolerance (Source: Lay and Anderson)
- 42. Anti fungal plant defensins Morphogenic defensins Reduced hyphal elongation with a concomitant increase in hyphal branching.Rs-AFP1, Rs-AFP2 Nonmorphogenic defensins Reduce the rate of hyphal elongation, but do not induce marked morphological distortions. Dm-AMP1, Dm-AMP2
- 43. Monoterpenes support SAR within and among plants Role of volatile organic compounds in SAR Arabidopsis thaliana Bicyclic monoterpenes α-pinene and β-pinene ROS Azelaic acid Info chemicals in plant-to-plant signaling Allowing defense signal propagation between neighboring plants. (Rieldmeier et al., 2017)
- 47. Long lasting immunity Long lasting immunity Long-lasting resistance to secondary pathogen attack Activate signaling components Secondary pathogen attack Require pathogen or chemical challenge for activation Transcriptionally active NPR1, MPK3, MPK6 Priming (Spoel and Dong, 2012)
- 48. Systemic wounding Response (SWR) Plants produce jasmonic acid and Methyl Jasmonate in response to particularly herbivory and wounding which build up in the damaged parts of the plant. MeJA is also a plant hormone involved in tendril (root) coiling and seed maturation. Act as signaling compounds for the production of phytoalexins. MeJA has been used to stimulate traumatic resin duct production in lodgepole pine trees. This can be used as a defense against many insect attackers as a type of vaccine. Oligogalacturonides (OGs) Fragments of pectin that activate plant innate immunity by functioning as damage-associated molecular patterns (DAMPs). Released at the site injury. .
- 50. (Ballare, 2014)
- 51. Systemic Gene Silencing (SGS)
- 52. SAR Benefits SA can stimulate ABA accumulation. Stomatal closure. Reduction of transpiration, thus allowing the storage of water in leaves for survival under drought conditions. (Miura et al., 2011).
- 53. Detoxification of ROS by SAR
- 55. Antioxidants and antioxidant enzymes acts as scavengers of ROS. Antioxidant enzymes. Ascorbate peroxidase(APX) Superoxide dismutase(SOD) Catalase(CAT) Peroxidase(POD) Effects of SA on mulberry plants
- 56. Critical limit of Micronutrients in soil for normal growth of Mulberry 56 Trace elements Conc. in ppm Fe 100 Mn 50 B 20 Zn 20 Cu 6 Mo 0.1 (Principles of Temperate sericulture (Afifa S. Kamili and M. A. Masodi)
- 57. Effect of SAR chemicals on per cent disease incidence and per cent disease control (45 and 70 days after pruning) under field conditions in mulberry. T1 = Salicylic acid; T2 = Isonicotinic acid; T3 = Calcium chloride; T4 = Ascorbic acid; T5 = Ethylene diamine tetra acetic acid; T6 = Sodium salicylate; T7 = β-amino butyric acid; T8 = Check (Carbendazim 50% WP) 0 10 20 30 40 50 60 70 80 Salicylic acid (1.5) Isonicotinic acid (2.0) Calcium chloride (10) Ascorbic acid (3.0) Ethylene diamine tetra acetic acid (1.0) Sodium Salicylate (2.0) β-amino butyric acid (2.0) Check (Carbendazim 50% WP) (0.5) Per cent disease control 45 days after pruning Per cent increase Per cent disease control 70 days after pruning Per cent increase Linear (Per cent disease control 70 days after pruning Per cent increase)
- 58. Effect of SAR chemicals on per cent disease control (40 and 70 days after pruning) under field conditions in mulberry. (Bandana Mazal, 2014) Treatments (Conc. mg/ml) Per cent disease control 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid (1.5) 48.64 52.2 Isonicotinic acid (2.0) 49.61 55.90 Calcium chloride (10) 40.24 44.71 Ascorbic acid (3.0) 36.25 44.34 Ethylene diamine tetra acetic acid (1.0) 26.08 41.56 Sodium Salicylate (2.0) 45.53 49.68 β-amino butyric acid (2.0) 69.24 74.57 Check (Carbendazim 50% WP) (0.5) 66.48 72.77
- 59. Effect of SAR chemicals on the percent disease intensity and disease control of the powdery mildew (Phyllactinia corylea) disease of mulberry leaves during 2010 and 2011 Kharief seasons. Chemical compounds PDI PDC Pooled 2010 2011 2010 2011 PDI PDC Ascorbic acid 6.84 4.18 68.23 72.26 5.51 70.14 Benzoic acid 993 6.08 54.80 59.65 7.90 57.22 Calcium chloride 7.32 5.14 66.00 65.89 6.23 65.94 Salicylic acid 6.17 3.65 71.34 75.77 4.91 73.55 EDTA 7.04 6.84 67.30 54.61 6.94 60.95 Check-I (Carbendazim 50 WP) 2.70 1.24 87.45 91.77 1.97 89.61 Check-II (Untreated) 21.53 15.07 - - 18.30 PDI =Percent disease intensity PDC = Percent disease control (A.G.Najar, 2010)
- 60. Effect of SAR chemicals on nitrogen content in mulberry leaves of (Morus spp.) Treatment 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid 32.07 38.09 Isonicotinic acid 32.70 42.86 Calcium chloride 27.04 27.78 Ascorbic acid 16.98 22.22 Ethylene diamine tetra acetic acid 10.06 10.32 Sodium salicylate 28.93 32.54 β-amino butyric acid 57.22 88.09 Check (Carbendazim 50% WP) 33.96 50.79
- 61. Effect of SAR chemicals on phosphorus content in the leaves of mulberry (Morus spp.) Treatment 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid 39.09 70.37 Isonicotinic acid 43.64 74.67 Calcium chloride 22.73 41.67 Ascorbic acid 7.27 27.04 Ethylene diamine tetra acetic acid 5.45 29.63 Sodium salicylate 35.91 46.30 β-amino butyric acid 68.18 98.15 Check (Carbendazim 50% WP) 62.27 88.89
- 62. Effect of SAR chemicals on potassium content in the leaves of mulberry (Morus spp.) Treatment 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid 14.10 38.51 Isonicotinic acid 16.67 40.86 Calcium chloride 10.45 29.69 Ascorbic acid 9.17 28.36 Ethylene diamine tetra acetic acid 8.72 22.89 Sodium salicylate 13.65 31.48 β-amino butyric acid 19.42 44.22 Check (Carbendazim 50% WP) 18.33 43.20
- 63. Treatments Moisture (%) 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid 8.76 16.24 Isonicotinic acid 10.72 18.00 Calcium chloride 7.0 13.30 Ascorbic acid 5.55 11.89 Ethylene diamine tetra acetic acid 4.45 7.07 Sodium salicylate 8.20 13.61 β-amino butyric acid 15.25 21.19 Check (Carbendazim 50% WP) 13.70 18.43 Effect of SAR chemicals on moisture content in the leaves of mulberry (Morus spp.)
- 64. Treatments Total chlorophyll (mg/g) 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid 46.31 107.62 Isonicotinic acid 51.15 116.10 Calcium chloride 34.22 83.61 Ascorbic acid 24.55 53.95 Ethylene diamine tetra acetic acid 8.95 34.32 Sodium salicylate 41.48 93.50 β-amino butyric acid 60.82 145.76 Check (Carbendazim 50% WP) 55.99 124.57 Effect of SAR chemicals on total chlorophyll content in the leaves of mulberry (Morus spp.)
- 65. Effect of SAR chemicals on carbohydrate content in the leaves of mulberry (Morus spp.) Treatment 45 days after pruning Per cent increase 70 days after pruning Per cent increase Salicylic acid 38.91 45.34 Isonicotinic acid 40.79 46.36 Calcium chloride 34.28 39.21 Ascorbic acid 22.22 29.68 Ethylene diamine tetra acetic acid 18.86 25.60 Sodium salicylate 36.63 42.96 β-amino butyric acid 41.27 50.44 Check (Carbendazim 50% WP) 41.11 48.40
- 66. SAR inducers against Nematode Reproduction Acibenzolar-s-methyl DL- α-amino-n-butyric acid (AABA) DL-β-amino-n-butyric acid (BABA) Gamma-amino-n-butyric acid (GABA) P-aminobenzoic acid (PABA) Riboflavin Salicylic acid (SA) Acibenzolar decreased R. reniformis egg production by 58% compared to the nontreated control. Acibenzolar, BABA, and riboflavin reduced M. javanica egg production by 60% to 64% compared control. Meloidogyne javanica and Rotylenchulus reniformis in pineapple (Chinnasri et al., 2006)
- 67. SAR in mulberry against multiple diseases Leaf spot (Cercospora moricola) Leaf rust (Cerotelium fici) Azotobacter chroococcum strain Azc-3, Bacillus megaterium strain Bm-1 and Pseudomonas fluorescens strain Psf-4. Plant activators, acetyl-salicylic acid (ASA), sodium salicylate (NaS) and 4-amino-n-butyric acid (ABA). Integration of Azc-3 + ASA provided greater suppression to multiple infections of brown leaf spot and leaf rust diseases during the entire growth period of mulberry plants. Thus combination of biological and chemical elicitors holds great promise to provide an effective ecofriendly alternative to the toxic chemical fungicides presently recommended for the control of brown leaf spot and leaf rust diseases in mulberry. (Mishera et al., 2008)
- 68. Conclusions Promotion of SAR inducers reduces adverse effect on environment, humans and silkworms as well. All SAR chemicals give good protection against the diseases and are almost equally effective as fungicides. There is a need to promote the use of these chemicals in order to make agricultural crops sustainable. Promotion of these chemicals exploits the own resistance of mulberry against pests and diseases. This is an ecofriendly approach of disease and pest management. Stable, low risk of pathogen populations of developing resistance to SAR.
- 69. EVERYTHING ELSE CAN WAIT BUT NOT AGRICULTURE (J. Nehru)