Here is an elaborated discussion on conventional and and biotechnological breeding techniques used for disease resistance in plants.
Conventional Breeding Techniques for Disease Resistance in Plants:
The conventional methods of breeding for disease resistance are essentially the same as those for other agronomic characters.
The following breeding methods have been commonly used:
(i) Selection,
(ii) Introduction,
(iii) Mutation and
(iv) Hybridization.
(i) Selection:
Selection of resistant plants from a commercial variety is the cheapest and the quickest method of developing a resistant variety. The method has been useful in many cases in the past, but it has only a limited usefulness at the present level of crop improvement. ‘Kufri Red’ potato is a disease resistant selection from Darjeeling ‘Red Round’. Other examples include resistance to curlytop in sugar-beets, to mildew and leaf spot in alfalfa, to cabbage yellows in cabbage and to Periconia root rot in sorghum.
Pusa Swani bhindi (A. esculentus) is a selection from a collection from Bihar; it is comparatively resistant to yellow mosaic under field conditions. Cotton (Gossypium hirsutum) variety MCU 1 (Madras Combodia Uganda 1) was selected from the variety Coimbatore 4 (Co 4) for resistance to black-arm; it has an acceptable level of resistance under field conditions.
(ii) Introduction:
Resistant varieties may be introduced for cultivation in a new area. This offers a relatively simple and quick means of obtaining resistant varieties. Introduction is quick, but has certain limitations. Firstly, the introduced varieties may not perform well in new area. Secondly, they may become susceptible to the concerned disease in the new environment, and thirdly, they may be susceptible to the concerned disease to other diseases common in the new area.
For example, Kenya-wheats (Triticum aestivum) introduced in India were rust resistant, but they were highly susceptible to loose smut. Similarly, the American cotton (G. hirsutum) varieties introduced in India were susceptible to red blight.
Introductions have served as a useful method of disease control. For example, Ridley wheat introduced from Australia has been useful as a rust resistant variety. Early varieties of groundnut (A. hypogaea) introduced from U.S.A. have been resistant to leaf spot or tikka disease (caused by Cercospora arachidicola). Kalyan Sona and Sonalika wheat varieties originated from the segregating materials introduced from CIMMYT, Mexico, and were rust resistant.
Introductions also serve as sources of resistance in breeding programmes. For example, African bajra (Pennisetum americanum) introductions have been used for developing downy mildew resistant male sterile lines (Tift 23A cytoplasm) for use in hybrid bajra production. This has been a crucial development in the hybrid bajra programme since the original male sterile lines Tift 23A and 23D2A were extremely susceptible to downy mildew.
(iii) Mutation:
‘Mutants’ are those plants whose genetic characters have changed as a result of treatment with chemicals or physical effects (UV-light, X-rays, etc.) to such an extent that they become stable and transferable to progenies. The mutants possessing required degree of resistance to particular disease(s) and positive commercial value are selected, multiplied, and finally supplied to the farmers.
(iv) Hybridization:
Hybridization is the most common method of breeding for disease resistance.
Hybridization serves the following two chief purposes:
(i) Transfer of disease resistance from an agronomically undesirable variety to a susceptible but otherwise desirable variety (by backcross method), and
(ii) Combining disease resistance and some other desirable characters of one variety with the superior characteristics of another variety (by pedigree method).
In the first case (backcross method), the new variety is agronomically the same as the susceptible variety, but is disease resistant. In the second case (pedigree method), on the other hand, the new variety is expected to be superior to both the parents in agronomic characteristics and at the same time would be disease resistant.
In both the cases, one parent is selected for disease resistance; it should have a high intensity of resistance to as many races of the pathogen as possible, and the resistance should be governed by few oligogenes. When the resistant variety is un-adapted and agronomically undesirable, backcross method is the obvious choice. But when the resistant variety is well adapted and has some other desirable features as well, the pedigree method of breeding is preferred.
a. Pedigree Method:
The pedigree method is quite suited for breeding for horizontal or polygenic resistance as backcross method, in such cases, is of limited value. Pedigree method for breeding of disease resistance is not materially different from the method used for other quantitative traits, e.g., yield.
In breeding for disease resistance, artificial disease epidemics are generally produced to help in the selection for disease resistance. Therefore, we shall not examine the pedigree method of breeding for disease resistance in any detail.
A vast majority of disease resistant commercial varieties have been developed through the pedigree method of breeding, e.g., Kalyan Sona, Sonalika, Malviya 12, Malviya 37, Malviya 206, Malviya 234, and many other wheat varieties. G. hirsutum (American cotton) variety Laxmi, resistant to red leaf blight, was developed through pedigree method from the cross between susceptible parent Gadag 1 and the resistant parent Coimbatore Combodia 2.
b. Backcross Method:
The backcross method is useful in transferring genes for resistance from a variety that is undesirable in agronomic characteristics to a susceptible variety, which is widely adapted and is agronomically highly desirable. The backcross programme would differ depending upon the allelic relationship of the resistance gene, i.e., whether it is recessive or dominant to the allele for susceptibility.
Generally, 5-6 backcrosses are made, a selection for the plant type of the recurrent parent (the susceptible variety) during backcrossing, particularly in BC2 and BC3, is effective in making the backcross progeny resemble the recurrent parent rather rapidly. At the end of the backcross programme, the progeny are selfed and resistant plants are selected.
Progenies derived from different resistant plants that are identical in agronomic characteristics are usually bulked to produce the new disease resistant variety. The new variety would be almost identical to the recurrent parent, except for the disease resistance; hence extensive yield trials are usually not required before its release for commercial cultivation.
Biotechnological Breeding Techniques for Disease Resistance in Plants:
Biotechnological advances in the field of tissue culture technology and recombinant DNA technology (genetic engineering) have opened a whole new array of possibilities to raise resistance in hosts to diseases.
I. Tissue Culture Technology:
Clonal propagation, the tissue culture technique in which vegetative parts (shoot apex or auxiliary bud tissues) are used to produce plants that are genetically identical to their parents (i.e. the clones), is particularly useful in obtaining disease resistant plants. Clonal propagation facilitates the rapid propagation of plants with exceptional (resistant) genotypes, especially in those crops not easily propagated by seed (e.g., apples, bananas, cassava, potatoes, strawberries, sugarcane, etc.
i. Disease Resistant Mutants from Plant Cell Cultures:
Plants regenerated from callus, single cells, or protoplasts often show considerable variability (somaclonal variation) and are called somaclonal variants. Much of the somachonal variants are useless with disease resistance view point, few are useful.
Disease resistant somaclonal variants are often obtained in the following two ways:
a. Plants regenerated from cultured cells or their progeny are subjected to disease test and resistant plants are isolated (screening). For instance, when plants were regenerated from leaf protoplasts of a potato variety susceptible to both early blight (Alternaria solani) and late blight (Phytophthora infestans), some of the clones (5 of 500) proved resistant to early blight and some (20 to 800) proved resistant to late blight. Similarly, plants regenerated from sugarcane exhibited increased resistance to diseases caused by Ustilago scitaminea (whip smut) and Cochliobolus.
b. Cultured cells are selected for resistance to the toxin or culture filtrate produced by the pathogen and plants are regenerated from the selected cells (cell selection). This strategy is most likely to be successful in cases where the toxin is involved in disease development.
ii. Resistant Diploids from Haploid Plants:
Haploid (N) plants can be developed by inducing immature pollen cells (microspores) and, sometimes, megaspores of many plants. These haploids possess single copies of each gene (alleles) in all sorts of combinations.
Most highly resistant haploids can be sorted out from these haploids by vegetative propagation and proper screening. The selected haploids can be subsequently treated with colchicine, which induces diploidization of nuclei, i.e., doubling the number of chromosomes and production of diploid plants homozygous for all genes, including genes for resistance.
iii. Increase in Disease Resistance by Protoplast Fusion:
When protoplasts either from closely related or even from unrelated plants are fused under proper conditions, the fusion products may give rise to hybrid cells containing cytoplam and nuclei (chromosomes) of both protoplasts, and cybrid cells containing the cytoplasm of one protoplast and the nuclei (chromosomes) of the other protoplast. Hybrid cells of unrelated protoplasts generally abort, or may give rise to calluses, but they always fail to regenerate plants.
In hybrid cells from closely related protoplasts and cybrid cells, one or a few chromosomes of one protoplast may get incorporated in the genome of the other protoplast. In this way, plants with new characteristics can be regenerated from the products of protoplast fusion.
Protoplast fusion is particularly useful between protoplasts of different, highly resistant haploid lines of the same variety or species, and it can result in diploid plants that combine the resistance genes of two highly resistant haploid lines.
II. Recombinant DNA Technology (Genetic Engineering):
Genes expected to confer disease resistance are isolated, cloned, and transferred into the crop in question. There are now numerous examples of plants engineered (transgenic plants) for improved resistance to pathogens.
In case of bacterial and fungal pathogens, resistance has been sought by expression of the following transgenes:
(i) Genes encoding insensitive target enzymes,
(ii) Genes specifying toxin inactivation,
(iii) Expression of antibacterial peptides,
(iv) Expression of bacterial lysozymes,
(v) Genes specifying artificially programmed cell death (in items 1-5, transgenes are from non-plant sources),
(vi) Expression of heterologous phytoalexins,
(vii) Genes encoding ribosome inactivating proteins,
(viii) Expression of heterologous thionins,
(ix) Ectopic expression of pathogenesis related proteins and
(x) Ectopic expression of chitinases (items 6-10 use plant genes).
In almost all the approaches, transgenic plants showed increased resistance to the concerned diseases. In case of viral pathogens, several transgenes have been evaluated, viz., virus coat protein gene, DNA copy of viral satellite RNA, defective viral genome, antisense constructs of critical viral genes, and ribozymes’. Viral coat protein gene approach seems to be the most successful. A virus resistant transgenic variety of squash is in commercial cultivation in U.S.A.
However, the large majorities of the transgenes are still at an experimental stage and are under trials in model crops such as tobacco and potato.
i. Genetic Engineering and Artificially Programmed Cell Death:
The strategy of artificially programmed cell death has been designed to mimic hypersensitive response. An artificially programmed cell death is brought about by endogenous gene action, particularly in response to some specific stimulus, e.g., the elicitor specified by avr genes in the case of hypersensitive response.
However, hypersensitive response depends on specific pairs of avr and R genes; therefore, each such pair specifies resistance to a single race of a pathogen and is not of general applicability. In contrast, the artificially programmed cell death is so designed as to cover all the races of a pathogen and possibly, more than one pathogen as well.
There are two schemes for artificial cell death, viz.:
(a) Two-component and
(b) Single-component systems.
In the two-component system, two precisely matched transgenes are expressed in the same cell. One transgene is an avr gene, say avr9, driven by a promoter inducible by a nonspecific elicitor produced during/by pathogen invasion. The other transgene is the R gene corresponding to the avr gene used, i.e., cf9 in this case; this transgene is driven by a constitutive promoter.
The expression of avr gene must be precisely regulated – it must be expressed immediately following pathogen attack but only in the infected cells. Expression of the avr gene would produce the elicitor that would be recognized by the corresponding R gene product; this recognition will initiate and culminate in hypersensitive response. This scheme should be applicable to any plant-pathogen combination, provided that the avr gene is driven by the appropriate promoter. So far, this scheme has not been tested.
The single-component system is based on the expression of a toxic polypeptide in response to pathogen infection. The transgenes usable in this scheme may be those that encode toxins, ribonucleases, or other enzymes whose products are toxic to plant cells. The Barnase gene from B. amyloliquefaciens was placed under the control of infection-specific promoter prp 1- 1 and was transferred into potato.
Transgenic potatoes showed effective control of Phytophthora infestans. Promoter prp 1-1 specifies the expression of barnase gene in such cells that are infected by a fungal pathogen. Synthesis of Barnase protein, an RNAase, in such cells leads to their death; the pathogen would die along with the dying host cells. Obviously, the strategy of artificially programmed cell death will not be effective against facultative parasites.
ii. Genetic Engineering and Pathogen-Derived Resistance (PDR):
Pathogen-derived resistance (PDR) refers to the resistance based on genes taken from the genome of virus pathogen and engineered into the host. In principle, it has been realized that any part of a plant viral genome can potentially give rise to pathogen-derived resistance (PDR).
However, some important features of PDR are:
(a) The level of resistance is often variable because it depends on the viral sequence used and the behaviour of plants to be transformed. In certain cases, one finds only sight attenuation of symptoms, while in others almost complete resistance shown by the transformed plant to infection, and
(b) Pathogen-derived resistance in often quite specific for the viral pathogen from which the transgene was derived.
The exact mechanism of pathogen-derived resistance is still uncertain. In certain cases, it appears that the expression of protein is necessary for effective resistance, while in other cases the presence of viral genome alone serves the purpose. Scientists have achieved some limited success in expressing an antisense version of viral RNA. In this case, the viral RNA binds to the complementary sense strand and interferes with replication.
However, different types of PDR together provide a promising new way to control infections by viral pathogens. The full potential of PDR, and its prolonged durability in the field, has yet to be evaluated, but its pioneering results are encouraging.
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