In this article we will discuss about how do plants protect and defend themselves from pathogens and diseases.
Introduction to Pathogens:
Once a pathogen has arrived in the vicinity of a potential host plant or, as may happen in the case of soil-borne pathogens, a plant root has arrived in the vicinity of a pathogen, subsequent events depend on the production and perception of signals by both partners.
First we shall be concerned with those factors external to the plant that influence the behaviour of propagules of the pathogen and then the host and pathogen factors that are involved in the mechanisms of penetration.
Since some of these, such as degradative enzymes, are also involved in further colonization of the host after penetration, this aspect of parasitism will also be considered. In soil, pathogens may be influenced by compounds exuded from the host root.
Motile stages may be attracted or repelled and the germination of sessile propagules stimulated or inhibited. Air-borne pathogens generally rely upon large populations of propagules to ensure that at least some of them alight on a suitable host.
At this point adhesion is a necessity to prevent the propagule being washed off the plant and, for at least one fungal pathogen, adhesion has been established as a prerequisite for germination.
Following adhesion, germination, which may be under the control of topological or chemical signals from the host, occurs and in some instances such signals lead to the differentiation of infection structures. These, too, require firm anchoring to the surface of the plant if any mechanical force is to be exerted.
Some pathogens enter their hosts via natural openings such as stomata, nectaries, hydathodes or lenticels while others require wounds, which may be made by physical phenomena such as wind or hail or by biological agents such as vectors or herbivores.
They therefore require neither mechanical force nor enzymes to establish their initial beachhead in the host. Others use mechanical force to affect entry but often these rely upon the secretion of enzymes as well.
Such enzymes include lipases and cutinases for breaching the wax and cuticle of aerial parts of the plant as well as enzymes for degrading cell-wall constituents such as pectic substances, cellulose and lignin.
A significant number of pathogens live extra-cellularly in their hosts and some are confined to conducting tissue. These would seem to have only limited requirements for degradative enzymes.
Biotrophs often feed through haustoria, which penetrate the host cell wall, almost certainly through the agency of degradative enzymes, and invaginate but do not penetrate the host plasma membrane. For necrotrophs the role of degradative enzymes seems clear.
They are required not only for penetration and colonization of plant tissue but also to reduce the high molecular weight components of these tissues to products which they can metabolize.
In the case of soft rotting organisms this often results in the ‘mushy’ symptoms that give these diseases their name. Despite the seemingly obvious necessity of enzymes for pathogenicity or virulence, unequivocal demonstration of such roles has been difficult to achieve owing to the numbers and diversity of isozymes produced by pathogens and subtleties in their regulation.
Colonization of the host by viruses is a special case as they lack enzymes with which to degrade plant cell walls but spread from cell to cell through plasmodesmata and then, in the case of systemic infections, by way of the phloem or xylem.
Since many pathogens need to penetrate the outer coverings of plants and plant cell walls in order to establish a parasitic relationship with their hosts, the characteristic of these will be described first.
Aerial parts of plants are generally covered with a cuticle consisting of an outer, thin layer of wax and an inner layer of cutin. The outer waxy layers are usually complex and a stereoscan electron microscopy survey of 13 000 species representing all the major groups of seed plants has classified them into 23 types.
This physical diversity is also paralleled by their chemical diversity. For example – Shepherd and co-workers (1999) found that the epicuticular waxes of red raspberry consisted of a complex mixture of free primary alcohols and their acetates, secondary alcohols, ketones, terpenoids including squalene, phytosterols, tocopherol, amyrins, alkanes and long chain alkyl and terpenyl esters.
Similarly, Jetter and Riederer (2000) examined the rodlet-shaped wax crystals of the fern, Osmunda regalis, and found 139 compounds belonging to 14 homologous series. These included alkanes alkyl esters, primary alcohols, secondary alcohols, ketones, aldehydes, fatty acids and b-sitosterol.
Clearly surface waxes are far from simple and potentially contain an abundance of physical and chemical cues, which may influence the pre-penetration stages of organisms seeking entry into plants.
Cutin, which is found under the wax surface, is an amorphous substance composed of hydroxylated and epoxylated fatty acids with either 16 or 18 carbon atoms and these are linked to each other by means of ester bonds.
Underground parts of plants are generally covered with suberin. This is more complex than cutin and consists of alcohols and monobasic acids with 18 to 30 carbon atoms as well as hydroxylated or dibasic acids with 14 to 20 carbon atoms together with phenolic components.
The phenolics are unique and distinct from lignin, consisting primarily of hydroxycinnamates such as feruloyltyramine. Comparatively recently, evidence has been presented for the old hypothesis that glycerol is an important monomer of suberin. Suberization is also an active mechanism by which plants defend themselves from attack by pathogens.
The cell walls underlying cutin or suberin consist of two parts, the outer primary wall and the inner secondary wall. Both are composed of a crystalline microfibrillar phase of cellulose, a b-1, 4 linked glucan, embedded in an amorphous phase.
The micro-fibrils are about 30nm in diameter and consist of aggregates of 20-70 linear chains of cellulose, which are held together by hydrogen bonds between the sugar OH groups.
The incidence of micro-fibrils in primary walls is sparse and this comparative scarcity allows the cell to grow. Once growth has ceased, the secondary wall is laid down which contains an orderly arrangement of cellulose micro-fibrils as well as hemicelluloses.
The latter are a heterogeneous group of saccharide polymers which may be extracted with alkali and, on hydrolysis to their monomers, yield glucose, xylose and arabinose.
In monocotyledonous plants arabinoxylan is not only the predominant hemicellulose but may account for up to 60 per cent of the total wall carbohydrate. It is composed of a b- 1, 4-linked D-xylopyranosyl backbone which is frequently substituted at the O-2 and O-3 positions by various mono- or oligo-saccharide units, consisting mainly of arabinosyl-, xylosyl- and/or glucosyluronic acids. The middle lamella lies between cells and is composed largely of pectic substances.
The predominant motif is a linear chain of galacturonic acid residues which are a-1, 4 linked. Where this is the major constituent the substrate is referred to as polygalacturonate or pectate but often the carboxyl groups of the galacturonate residues are esterified with methanol and then the term pectin is used.
The carboxyl groups of polygalacturonate form salts with the divalent cation, calcium, thus linking neighbouring chains. This gives rise to the gel properties of pectin and provides the cohesive element that cements cells together.
The ‘smooth’ regions of pectate or pectin may be interspersed by the occasional rhamnose residue and these may be substituted with oligosaccharides composed of neutral sugars such as arabinose, galactose and xylose giving rise to so-called ‘hairy’ regions!
In addition, some of the galacturonate residues are acetylated at the C-2 or C-3 position. Arabinose and galactose are also prominent constituents of the middle lamella, forming the polymers, arabinan and galactan, respectively and are linked to the rhamnogalacturonan backbone.
Lignin is another important component and constitutes between 15 and 35 per cent of the dry weight of woody tissues. It is a heterogeneous polymer formed by the polymerization of up to three components – coumaryl, coniferyl and sinapyl alcohols.
Proteins, which may have structural or enzymatic functions, can account for as much as 15 per cent of the cell wall. Most of those that have been characterized are from dicotyledonous species.
They consist of extensins, which are hydroxyproline-rich glycoproteins (HRGPs), glycine-rich proteins (GRPs), proline-rich proteins (PRPs, an abbreviation not used to describe such proteins in this text in order to avoid confusion with pathogenesis-related proteins), arabino-galactan proteins (AGPs) and solanaceous lectins.
Variants of HRGPs are found in monocotyledonous species, which contain threonine and histidine hydroxyproline-rich glycoproteins (THRGPs and HHRGPs, respectively).
Although once the secondary cell wall has been laid down its apparently rigid structure would appear to preclude any further change it is, nevertheless, dynamic and responds to stimuli such as challenge with pathogenic organisms.
Several taxa of soil-borne pathogens have motile stages. These include Chytridiomycetes, nematodes, Oomycetes, Plasmodiophoromycetes and bacteria. Since plants release as much as 20 per cent of their photosynthate into the rhizosphere it is not surprising that some of the compounds found in their exudates influence the motility of these pathogens. Furthermore, evidence is accumulating that root cap border cells have important roles to play in determining whether or not the roots themselves become infected by such organisms.
Border cells originate from the root cap meristem and are attached to the root periphery by a water-soluble polysaccharide matrix, but their middle lamellae are solubilized by pectolytic enzymes in the cell wall. As a result, the cells immediately disperse when root tips are placed in water. Border cells affect both bacterial and fungal plant pathogens by chemo-attraction or repulsion and there is some evidence that they may act as decoys of pathogenic fungi, thus protecting the root from infection.
For example – Hawes and co-workers (1998) found that when pea roots were uniformly infected with spores of a virulent strain of the fungus, Nectria haematococca, hyphal growth was confined to the apex forming a mantle, which, on placing in water, fell off as a unit.
The mantle supported copious growth of the fungus but at the surface of the root only newly formed border cells and not fungal hyphae were apparent.
Zoospores of Oomycetes frequently accumulate at the zone of elongation of roots and it was initially thought that there were generally no appreciable differences in accumulation between plants that are resistant and those that are susceptible.
However, more recent work has refuted this view for a number of plant and pathogen combinations. For example, Mitchell and Deacon (1986) investigated four species of Pythium, P. graminicola and P. arrhenomanes, which infect only grass species and P. aphanidermatum and P. ultimum which have broad host ranges.
When grass roots or roots of dicotyledonous plants were placed in zoospore suspensions, larger numbers of the two graminicolous species accumulated on grass roots than on roots of dicotyledonous plants whereas there was no differential effect on the species with broad host ranges.
More specifically, zoospores of Phytophthora megasperma f. sp. sojae which infect soybean were strongly attracted to two flavonoids produced by the plant, daidzein and genistein – concentrations as low as 0.01 mM being effective.
In contrast, strains of P. megasperma that were infective for lucerne and Douglas fir did not respond to these compounds. In further work, Tyler and co-workers (1996) investigated 59 compounds which were structurally related to genistein and daidzein and found that 43 elicited some response.
In particular, the possession of a hydroxyl group at the 4′ and 7 positions of the isoflavone molecule or comparable positions of related molecules was necessary for high activity. Wide differences in sensitivity of strains of the fungus were noted and also some analogues were repellent.
Using a different Oomycete, Aphanomyces cochlioides, Horio and co-workers (1992) found that another flavonoid, cochliophilin A-, from spinach is an effective attractant of zoospores at 0.001 mM.
The volatile compound, isovaleraldehyde, is effective as a chemoattractant for zoospores of Phytophthora palmivora and P. cinnamomi and the latter Oomycete was also responsive to several amino acids, alcohols and phenolics.
Additionally, Cahill and Hardham (1994b) demonstrated that zoospores of P. cinnamomi showed strong electrotaxis towards a positively charged nylon membrane. Exploiting both chemotaxis and electrotaxis as well as a specific monoclonal antibody they were able to detect as few as 40 zoospores/ml in an assay time of less than 40 min.
Zoospores of Pythium dissotocum accumulate on border cells of cotton and penetrate them within minutes, although the chemo-attractant has not been identified.
Acetosyringone, which is secreted by tobacco and other plants in response to wounding, attracts Agrobacterium tumefaciens, the cause of crown gall. The same compound also activates plasmid- encoded virulence genes, which are involved in the transfer of DNA from the bacterium to the host.
After a zoospore has arrived at the surface of the host plant it normally encysts, the cysts then germinate and the resulting germ tubes usually grow towards the root, i.e. they show tropism. Jones, Donaldson and Deacon (1991) found that only glutamic and aspartic acids elicited zoospore taxis, encystment, cyst germination and tropism of germ tubes of Pythium aphanidermatum although a number of other compounds elicited one or more of these four responses.
The concentrations of the two acids used was quite high (25 mM), but it does not appear to be known whether such concentrations occur in the rhizospheres of susceptible plants or whether a synergistic mixture of these and other compounds is responsible for the sequence of the four events leading to infection.
Donaldson and Deacon (1993), investigating encystment of three species of Pythium, found that only P. catenulatum responded to fucosylated xyloglucan, fucoidan and a methylglucuroxylan, only P. aphanidermatum responded to arabinoxylan and only P. dissotocum responded to a non- fucosylated xyloglucan.
However, all three species encysted in response to gum arabic (100 mg/ml), sodium alginate (250-500mg/ml) or polygalacturonate (500-1000mg/ml). Morris, Bone and Tyler (1998) extended their work with Phytophthora sojae and flavonoids and found that daidzein and genistein, besides acting as chemo attractants of zoospores of the fungus, also exerted a chemotropic effect on hyphae from geminating cysts similar to that of roots of soybean.
Clearly, our knowledge of host factors that are interpreted as signals by pathogens and cause them to aggregate, encyst, germinate and grow towards the host is fragmentary.
Some fungi and bacteria enter their hosts passively through stomata, nectaries, hydathodes or lenticels. In particular, as in the case of wildfire of tobacco, bacteria notoriously enter plants through stomata by the agency of wind-driven rain.
Similarly, nectaries provide the point of entry of the fire blight pathogen Erwinia amylovora. Although stomata or trichomes provide a means of entry into tomato leaves by Clavibacter michiganensis subsp. michiganensis, it was not clear that infection by these routes could cause the marginal necrosis of leaves, sometimes known as ‘firing’, so often seen.
Carlton, Braun and Gleason (1998) found that hydathodes were the route by which the bacterium entered and caused such lesions. Under high humidity, hydathodes exude droplets of guttation fluid but these are withdrawn when conditions become dryer.
In these circumstances, should the guttation droplets become contaminated with the bacterium, infection leading to ‘firing’ occurs. Pathogens, which enter through lenticels, are generally those that require wounds, lenticels being of secondary importance as a means of entry.