How plants defend themselves against pathogens?

Plant Defense Mechanisms

Plants have evolved a multi-tiered defense system: 

• Pre-existing defense (passive defense)
• Pre-existing structural defense
• Pre-existing biochemical defense
• Post-infection or induced defense (active defense)
• Induced structural defense
• Induced biochemical defense 

A. Pre-existing Defenses

These are physical and chemical barriers present before infection.

• Pre-existing Structural Defenses: These include physical barriers that prevent entry.

• Cuticle: A thick, waxy cuticle repels water. Since many pathogens require moisture accumulation in the spore microclimate they cannot infect.

• Hairy Surface: A dense layer of trichomes (hairs) can deter insects that transmit pathogens. It also has water repellent effect.

• Stomata: The size and position of stomata can influence a pathogen's ability to enter.

• Cork Layers: A thick layer of cork cells can prevent the spread of pathogens.

• Sclerenchymatous tissue in the epidermis and hypodermis (make direct penetration difficult or impossible).

• Pre-existing Biochemical Defenses:

  • Exudates: 

• Some plants release antifungal substances from their roots or leaves. 
• For example, resistant varieties of onions (red scale onion) release protocatechuic acid and catechol, which inhibit the germination of fungal spores. The susceptible varieties (white scale onion) cannot produce these exudates.
• Fungitoxic exudates on the leaves tomato and sugar beet, inhibit the germination of spores of fungi Botrytis and Cercospora, respectively.

• Phytoanticipins: 

• Phytoanticipins are pre-formed antimicrobial compounds that are present in plants even before a pathogen has attacked. 
• Unlike phytoalexins, which are synthesized after infection, phytoanticipins are a part of the plant's constitutive defense system. 
• They are typically stored in an inactive form, such as a glycoside, and are activated by enzymes when the plant cell is damaged during an attack.
• These compounds can include:
• Phenolic compounds, tannins, and certain fatty acid-like compounds, such as dienes, that are found in high concentrations in young fruits, leaves, or seeds. They are potent inhibitors of various enzymes, including the pectolytic-macerating enzymes used by pathogens to break down host tissue.
• Saponins, which are preformed compounds with antifungal membranolytic activity. They work by disrupting the membranes of fungal pathogens, effectively excluding pathogens that lack the saponinase enzyme needed to neutralize them. Examples of saponins include tomatine in tomatoes and avenacin in oats.
• Glucosinolates: Found in plants of the mustard family (Brassicaceae), glucosinolates are hydrolyzed by an enzyme called myrosinase upon tissue damage. This reaction produces toxic compounds like isothiocyanates, which are effective against a wide range of pathogens and pests.

• Antimicrobial proteins: 

Plant proteins play a significant role in pre-existing defense against pathogens by performing several key functions:

• Inhibiting pathogen enzymes: These proteins can block pathogen proteinases and other hydrolytic enzymes that are used to break down the host cell wall.
• Damaging pathogen cell components: Some plant proteins can increase the permeability of fungal plasma membranes, causing them to leak, or inactivate foreign ribosomes, halting pathogen protein synthesis.
• Breaking down pathogen cell walls: Plant cells, particularly on their surfaces, contain hydrolytic enzymes like glucanases and chitinases. These enzymes can break down the cell wall components of fungi, thereby contributing to resistance.

Examples of these defensive proteins include:

• Phytocystatins: A family of low-molecular-weight proteins that inhibit cysteine proteinases, which are carried in the digestive systems of nematodes.
• Lectins: Proteins that bind to specific sugars on the surface of fungi, causing cell lysis and inhibiting their growth. 

• Defense Through Lack of Essential Factors:

Plants can naturally resist pathogens by lacking certain essential factors that the pathogen requires for a successful attack.

This can occur in several ways:

1. Lack of an essential nutrient: Some pathogens have very specific nutritional requirements. If a host plant lacks a particular nutrient that a pathogen needs to grow and reproduce, the pathogen will be unable to cause disease.
2. Lack of a specific host-recognition site: Many pathogens need to bind to a specific receptor on the surface of a host cell to initiate infection. If the host plant lacks this specific binding site, the pathogen cannot enter the cell, and the infection is aborted. 
3. Lack of a host-specific toxin receptor: Host-specific toxins require a specific receptor on the host cell to cause damage. If the host plant lacks this receptor, it becomes resistant to the toxin, and the pathogen is unable to cause the disease, even if it produces the toxin. For instance, some oat varieties are resistant to the victorin toxin because they lack the specific receptor that the toxin targets.

B. Induced (Post-Infection) Defenses

These defenses are activated after a pathogen attack.

• Recognition: 

• PAMP triggered immunity: Plants recognize pathogens by detecting PAMPs (pathogen-associated molecular patterns) via specialized PRRs (pattern recognition receptors). This initial recognition triggers a defense response.
• Effector triggered immunity: A host plant's R-gene product directly or indirectly recognizes the product of a pathogen's Avr-gene product (an effector molecule). This recognition event typically triggers a strong, localized defense response.

• Cytoplasmic Defense Reaction

In non-compatible interaction, the cytoplasm and nucleus enlarge. The cytoplasm becomes granular and dense, and various particles or structures appear in it. Finally, the mycelium of the pathogen disintegrates, and the invasion stops.

• Cell Wall Defense

The plant's cell wall is a dynamic structure that can be actively modified to strengthen its defenses and prevent the pathogen's spread.

• Papillae Formation: The host cell rapidly deposits new material, such as callose, lignin, and phenolics, to form a localized cell wall apposition called a papilla at the site of attempted penetration. This acts as a physical barrier to block the pathogen's entry.

• Strengthening of Existing Cell Walls: The plant can reinforce its existing cell walls by depositing additional layers of lignin, suberin, or other structural polymers. This makes the cell wall tougher and more difficult for the pathogen to penetrate or degrade.

• Restructuring of cell wall at the site of infection: The outer layer of the cell wall of parenchyma cells coming in contact with incompatible bacteria swells and produces an amorphous, fibrillar material that surrounds and traps the bacteria and prevents them from multiplying. 

•  Histological Defense Structures:

• Cork Layers

Cork layers are new layers of tissue that a plant forms around an infection site. This process effectively isolates the pathogen by creating a physical barrier, cutting off its access to healthy tissue and nutrients, and preventing it from spreading further into the plant.

• Tyloses

Tyloses are overgrowths of parenchyma cells that push into the xylem vessels, which are responsible for water transport. When a vascular pathogen enters the xylem, the plant responds by forming these balloon-like structures. The tyloses swell and block the vessel, physically stopping the pathogen's systemic movement through the plant's vascular system.

• Gum Deposition

Gum deposition is the process where a plant secretes gummy substances into and around infected cells. These gums form an impenetrable, sticky seal that encases the pathogen. This not only traps the pathogen but also limits its access to nutrients and water, effectively isolating it and preventing its spread.

• Abscission layer

It involves the controlled shedding of an infected part of the plant, such as a leaf or a fruit, to prevent the pathogen from spreading to the rest of the plant. These layers break down, effectively dissolving the connection to the main body of the plant. The infected part then falls off, taking the pathogen with it and leaving a clean, sealed wound that is resistant to further infection.

• Hypersensitive Response (HR)

It is a localized, induced cell death that occurs at the site of infection by a pathogen to limit its spread. It is an incompatible host-pathogen interaction.

The HR is a classic example of effector-triggered immunity (ETI), a key component of the gene-for-gene model of plant defense. This response is initiated when a plant's resistance (R) gene product recognizes a corresponding avirulence (Avr) gene product (or effector) from the pathogen.

The HR response involves a series of cascading biochemical and structural changes in the plant cell:

• Reactive Oxygen Species (ROS) Burst: One of the first events is a rapid and temporary generation of ROS, such as superoxide (O2−​), hydrogen peroxide (H2​O2​), and hydroxyl radicals (OH−). This leads to the hydroperoxidation of membrane phospholipids, causing the disruption of cell membranes and the eventual death of the cell.

• Ion Movement: There is a rapid increase in the movement of ions, particularly an efflux of K+ and H+ ions from the cell. This makes the cytoplasm more acidic and the extracellular space more alkaline.

• Cell Wall Strengthening: The plant strengthens its cell walls by cross-linking phenolic compounds with cell wall components, which leads to the formation of lignin-like substances. The cell wall also receives deposits of other defensive substances like callose and glycoproteins, making it harder for the pathogen to penetrate.

• Production of Antimicrobial Substances: The HR triggers the synthesis of various antimicrobial compounds:

  • Pathogenesis-Related (PR) Proteins: These include proteins like chitinases and β-1,3-glucanases that can break down the cell walls of fungi.

  • Phytoalexins: These are toxic, low-molecular-weight substances that are synthesized and accumulate in the plant after infection to inhibit pathogen growth.

  • Phenolic compounds: The oxidation of phenols by enzymes like polyphenol oxidases (PPO) and peroxidases creates highly toxic quinones, which further contribute to resistance.

By sacrificing a small number of cells, the plant effectively creates a barrier of dead tissue that confines the pathogen, preventing it from obtaining nutrients and spreading to other parts of the plant. 

Pathogenesis related proteins

Pathogenesis-related (PR) proteins are a diverse group of plant proteins that are synthesized and accumulate in plant cells in response to an attack by a pathogen. They are considered a key component of the plant's induced defense system.

PR proteins are either extremely acidic or extremely basic and therefore are highly soluble and reactive. 

These proteins have a wide range of functions, all aimed at protecting the plant from invaders:

• Enzymatic Activity: Many PR proteins are enzymes that can directly attack the pathogen's cell components. Examples include chitinases and β-1,3-glucanases, which break down chitin and β-1,3-glucans, two major structural components of fungal cell walls.

• Antimicrobial Properties: Some PR proteins, like certain defensins and thionins, have direct antimicrobial activity, disrupting pathogen membranes or interfering with their metabolism.

• Signaling: Other PR proteins may not directly harm the pathogen but instead act as signaling molecules to activate and amplify the plant's defense response throughout the plant, a process known as systemic acquired resistance (SAR).

PR proteins are classified into several families based on their structure and function. They are often produced in high concentrations in the area surrounding the infection site and are a crucial part of the plant's strategy to contain and eliminate pathogens.

Examples: 

•  PR1 proteins (antioomycete and antifungal) 
• PR2 (b-1,3-glucanases): breaks down chitin in the fungal cell walls 
• PR3 (chitinases): breaks down chitin in the fungal cell walls 
• PR4 proteins (antifungal) 
• PR6 (proteinase inhibitors) 
• Defensins: accumulate through ethylene and jasmonate pathway and have antimicrobial activity 
• Lipoxygenases: generate antimicrobial metabolites as well as secondary signal molecules such as jasmonic acid 
• Lysozymes: degrade glucosamine & muramic acid of bacterial cell wall 

Phytoalexins

Phytoalexins are low-molecular-weight, toxic antimicrobial substances that plants produce and accumulate rapidly in response to an attack by a pathogen or an environmental stressor. Unlike phytoanticipins, which are pre-formed, phytoalexins are synthesized de novo (from scratch) after the plant has recognized an invader.

Phytoalexins are produced by healthy cells adjacent to localized damaged and necrotic cells in response to materials diffusing from the damaged cells.  

Key characteristics of phytoalexins include:

• Induced Synthesis: They are not present in healthy, unchallenged plant tissues but are quickly produced at the infection site to limit the pathogen's growth and spread.

• Broad-Spectrum Toxicity: Phytoalexins are often toxic to a wide range of fungi and bacteria, but they are generally less toxic to the plant's own cells.

• Chemical Diversity: Phytoalexins are a chemically diverse group of compounds, including isoflavonoids, sesquiterpenoids, stilbenes, and polyacetylenes.

• Role in Defense: Their accumulation is a key component of the plant's hypersensitive response (HR) and other resistance mechanisms, as they effectively inhibit pathogen growth and development.

Some well-known examples of phytoalexins include:

• Rishitin in potatoes.
• Phaseollin in beans.
• Medicarpin in alfalfa.
• Resveratrol in grapes

Phytoanticipins vs Phytoalexins

Feature	Phytoanticipins	Phytoalexins
Timing of Production	Pre-formed; present in the plant before a pathogen attack.	Induced; synthesized and accumulated in response to a pathogen attack.
Origin	Part of the plant's constitutive defense system.	Part of the plant's post-infection induced defense response.
Function	Act as a pre-existing barrier, often stored in an inactive form and activated upon cell damage.	Directly inhibit pathogen growth and development at the infection site.
Examples	Saponins like tomatine (in tomatoes) and avenacin (in oats).	Terpenoids like rishitin (in potatoes) and isoflavonoids like phaseollin (in beans).
Role	Contributes to basic resistance by deterring non-adapted pathogens.	Contributes to gene-for-gene resistance and the hypersensitive response (HR).

 

How pathogens attack plants?

Host-Pathogen Interaction

The interaction between a plant host and a pathogen is a complex relationship determined by the pathogen's ability to infect and the host's ability to defend itself.

• Compatible Interaction: 

• This occurs when a pathogen successfully infects a susceptible host, leading to disease. 
• The pathogen can overcome the plant's defenses, grow, and reproduce. 
• Example: Infection of a susceptible wheat variety by the fungus Puccinia triticina, which causes leaf rust.

• Incompatible Interaction: 

• This is a resistant reaction where the pathogen is unable to establish a successful infection. 
• The plant's defenses are strong enough to prevent the pathogen from growing, and no disease symptoms develop. 
• Example: A resistant wheat variety may recognize the Puccinia triticina fungus and trigger a rapid defense response, preventing the infection from spreading.

Pathogen Attack Mechanisms

Pathogens employ a series of strategies to attack and infect their host, which can be broken down into three stages.

A. Pre-Penetration Stage

Pathogens navigate towards a host plant using various cues.

• Chemotaxis: Pathogens are attracted to specific chemicals secreted by the host plant. For example, sugars and amino acids released by a host can attract the zoospores of fungi like Pythium.

• Electrotaxis: Host plants generate a flow of electric currents from their cells, which can guide pathogens.

• Rheotaxis: Pathogens, particularly zoospores, can move through water currents, and if their speed matches the water flow, they can find and attack a host.

B. Penetration Stage

Once at the host surface, pathogens penetrate the plant through natural openings, wounds, or by direct penetration.

1. Natural Openings:

   • Stomata: Bacteria can swim in a film of water on the leaf surface and enter the stomatal cavity. Fungi, like those causing rust, can form a specialized structure called an appressorium that precisely aligns over the stoma before sending a hypha into the leaf.

   • Lenticels: These are pores on fruits, stems, and tubers that provide easy entry for pathogens, such as the bacterium Streptomyces scabies, which causes potato scab.

   • Hydathodes: These pores on leaf margins and tips secrete nutrient-rich guttation fluid, which pathogens like Xanthomonas campestris pv. campestris (black rot of cabbage) can use to enter the plant.

   • Nectaries: Nectar-secreting glands in flowers are exploited by pathogens like Erwinia amylovora (fire blight of apple and pear) for entry.

2. Wounds: 

• Injuries caused by weather, insects, or farming equipment create openings that bypass the plant's natural defenses. 
• Most viruses, for example, rely on wounds created by insect vectors for transmission.

       3. Direct Penetration: 

This method relies on both mechanical pressure and enzymatic degradation to break through the host's surface.

A. Cutinases: 

• The first line of defense a pathogen encounters is the cuticle, a waxy layer on the plant's surface.
• Pathogens secrete cutinases and non-specific esterases to break down cutin, the primary component of the cuticle, allowing the pathogen to reach the cell wall.
• Cutin esterase catalyzes the hydrolysis of ester bonds occurring between free hydroxyl and carboxyl groups of cutin.
• Carboxycutin peroxidase catalyzed the hydrolysis of peroxide groups of cutin. 

B. Pectinases: 

• The pathogen then targets the cell wall. 
• Pectinases are a class of enzymes that degrade pectic substances, which are key components of the middle lamella and primary cell wall. 
• This class includes:
• Pectinmethyl esterases (PME): 
• Causes hydrolysis of methyl ester groups of pectinic acid chain into methyl alcohol and pectic acid.
• Polygalacturonases (PG): 
• Called chain-splitting pectinase. 
• Splits the pectic chain by adding a molecule of water and breaking (hydrolyzing) the linkage between two galacturonan molecules.
• Pectin lyases (PL): 
• Splits the chain by removing a molecule of water from the linkage, thereby breaking it and releasing products with an unsaturated double bond.
  
  
  

C. Cellulases:

• After degrading the pectin, the pathogen uses four main enzymes to break down cellulose, the main structural component of the cell wall, into glucose:
• Cellulase C1: attacks native cellulose by cleaving cross linkages between chains 
• Cellulase C2:  also attacks native cellulose and breaks it into shorter chains
• Cellulase Cx: attacks these short chains and degrades them to the disaccharide cellobiose
• β-(1-4)-glucosidase: attacks cellobiose and degraded it into glucose

D. Hemicellulases:

• The degradation of hemicelluloses, another complex component of the cell wall, requires multiple enzymes such as xylanase, galactanase, and glucanase, which are secreted depending on the specific structure of the hemicellulose in the host plant.

E. Ligninases:

• Lignin is a tough, complex polymer that provides structural support to the plant. 
• Most pathogens cannot degrade it, but a group of fungi known as white rot fungi produce powerful enzymes called ligninases to break down and utilize lignin.

F. Proteases and Peptidases:

• These enzymes are produced to degrade the structural proteins and glycoproteins embedded in the plant's cell wall, further weakening the host's defenses.

C. Post-penetration Stage

This stage involves:

• Infection and colonization
• Growth and reproduction of the pathogen
• Development of symptoms
• Dissemination of pathogen for spread of infection

Pathogens can cause disease by producing toxins or manipulating the plant's own growth hormones.

1. Pathotoxins

Pathotoxins are proteinaceous, highly poisonous substances produced by pathogens, and they play a critical role in the development of plant diseases.

These toxins are effective even at very low concentrations and are known for their ability to bind tightly to specific sites within the plant cell.

Some pathotoxins are also unstable or react so quickly that they are difficult to isolate and study. 

These are poisonous substances that directly damage host cells. 

Their mechanism of action may involve:

• affecting the permeability of the cell membrane 
• inhibiting enzymes and subsequently interrupting the corresponding enzymatic reactions
• acting as antimetabolites

Toxins can be of two types: 

• Host specific toxin 
• Host non-specific toxin 

Host-Specific Toxins

• These toxins are highly selective, only causing disease in specific host plants that are susceptible to the pathogen. Their production is essential for the pathogen's ability to be virulent, and they are typically not produced outside host cells in laboratory cultures.
• Example: Victorin, T-toxin, etc.

 

Host Non-Specific Toxins

• In contrast to host-specific toxins, these toxins affect a wide range of plant species and are not necessary for the pathogen's virulence on a specific host. They can often be produced in vitro (in a lab setting) and can cause milder symptoms like chlorosis or necrosis. 
• Example: TAB-toxin, Tentoxin, etc.

 

 Comparison between host-specific and host-nonspecific pathotoxins

Feature	Host-Specific Toxins	Host Non-Specific Toxins
Specificity	Highly specific to a particular host plant.	Affect a wide range of host plants.
Virulence	Essential for the pathogen's ability to cause disease (virulence).	Not essential for the pathogen's virulence.
Production	Not typically produced in vitro (laboratory cultures).	Can be produced in vitro.
Toxicity	Highly toxic and effective at very low concentrations.	Generally, less toxic than host-specific toxins.
Examples	Victorin (Cochliobolus victoriae), T-toxin (Cochliobolus heterostrophus).	Tabtoxin (Pseudomonas syringae), Tentoxin (Alternaria tenuis).

2. Altered Growth Regulators

Pathogens can manipulate plant hormones for their own benefit.

• Auxins: Pathogens can increase auxin levels by inactivating the IAA oxidase enzyme. This can lead to the formation of tumors, galls, and hairy roots, such as the crown gall disease caused by Agrobacterium tumefaciens. Conversely, a decrease in auxin synthesis can result in stunted growth.

• Gibberellins: The fungus Gibberella fujikuroi produces excessive gibberellins, causing rice seedlings to grow tall and spindly, a disease known as "foolish seedling disease."

• Cytokinins: Some pathogens produce cytokinins, which can lead to excessive cell division and gall formation, or witches' broom symptoms. A reduction in cytokinin production can cause dwarfism. 

• Ethylene: Pathogens can alter ethylene levels, leading to premature yellowing (chlorosis) and senescence of leaves and fruits. Ex: Pseudomonas solanacearum infection in bananna 

 3. Polysaccharides in pathogenesis

Exo-polysaccharides appear to be necessary for several pathogens to cause disease symptoms by:

• being directly responsible
• indirectly facilitating pathogenesis by promoting colonization
• enhancing survival of the pathogen.

Example: Vascular wilts 

4. Suppressors of plant defense

Some pathogenic fungi produce substances called suppressors that act as pathogenicity factors by suppressing the expression of defense responses in the host plant. 

 Example: Puccinia graminis tritici