International Testing as a Means to Implement Durable Resistance: The Experience of CIMMYT with Wheat Rusts

The Gene‐for‐Gene Relationship in Plant–Parasite Interactions

Leaf or brown rust of wheat caused by Puccinia triticina (Pt) is one of the most damaging diseases globally. Considerable progress has been made to control leaf rust through crop protection chemicals and host plant resistance breeding in southern Africa. However, frequent changes in the pathogen population still present a major challenge to achieve durable resistance. Disease surveillance and monitoring of the pathogen have revealed the occurrence of similar races across the region, justifying the need for concerted efforts by countries in southern Africa to develop and deploy more efficient and sustainable strategies to manage the disease. Understanding the genetic variability and composition of Pt is a pre-requisite for cultivar release with appropriate resistance gene combinations for sustainable disease management. This review highlights the variability and distribution of the Pt population, and the current control strategies, challenges and future prospects of breeding wheat varieties with durable leaf rust resistance in southern Africa. The importance of regular, collaborative and efficient surveillance of the pathogen and germplasm development across southern Africa is discussed, coupled with the potential of using modern breeding technologies to produce wheat cultivars with durable resistance.

Rice blast disease caused by Pyricularia grisea, the anamorph of Magnaporthe grisea, is the main rice production constraint in Latin America. Development of resistant cultivars has been the preferred means of controlling this disease; however, blast resistance is defeated by the pathogen shortly after cultivar release, affecting both leaves and panicles of the plant and reducing yields severely. Major efforts are being made at CIAT to understand the high pathogen variation observed, often reported as the main cause of resistance breakdown. We have analyzed extensively the genetic structure of blast pathogen populations using MGR-DNA and rep-PCR fingerprinting techniques and studied the avirulence gene diversity using a set of rice differentials with known resistance genes. The blast pathogen in Colombia has been found to be mainly clonal exhibiting few genetic lineages. At present, there are three lineages predominating in the pathogen population and their frequencies depend upon the susceptibility and planted area of the commercial rice cultivars by farmers. In general, a single haplotype predominates within each lineage. These three genetic lineages exhibit broad spectrum of virulence and together defeat all known blast resistance genes. However, some resistance genes are effective against all members of a lineage, suggesting an association of avirulence genes and genetic groups in the pathogen. Avirulence genes vary in frequency in the pathogen population and some are highly frequent in several genetic lineages of the fungus. This suggests that these avirulence genes could play an important role in the pathogen or be associated with pathogenic fitness and then the corresponding resistance genes could be more relevant in breeding for durable resistance. Despite this high virulence diversity, breeders at CIAT have been able to develop durable blast resistant cultivars, indicating that combinations of these resistance genes may confer suitable and durable resistance to the pathogen. Evaluation and selection of breeding lines are routinely carried out under “hot spot” conditions favoring high pathogen pressure and diversity. Our studies are allowing us to identify and predict the durability of resistance gene combinations based on avirulence gene frequencies and the possible association of avirulence genes with pathogenic fitness. We have inferred the possible resistance genes present in our blast susceptible rice cultivars and initiated a backcrossing program to incorporate the desired resistance gene combinations into Latin American rice cultivars through marker assisted selection using mainly microsatellite and scar markers. Rice lines carrying the combination of the resistance genes Pi-1, Pi-2 and Pi-33 are then tested under controlled greenhouse conditions as well as our “hot spot” screening site using a spreader row technique to maintain a high and diverse population of the pathogen in the field. Resistant plants are then selected based on other desirable traits for their distribution to national programs in Latin America.

Durable resistance against fungal pathogens is highly valuable for disease management in agriculture. For its sustainable use, and to avoid pathogen adaptation, it is important to understand the underlying molecular mechanisms. Many studies on durable disease resistance in plants have focused exclusively on the host plant, whereas possible reactions and adaptations of pathogens exposed to this type of resistance have not been well researched. The wheat Lr34 gene, encoding a putative ABC‐transporter, provides broad‐spectrum and durable resistance against multiple fungal pathogens in wheat and is functional as a transgene in all major cereals. Lr34‐based resistance is partial, meaning pathogens can grow and reproduce to some degree on Lr34‐containing plants. Therefore, Lr34‐expressing plants are ideal for studying the response of pathogens to partial resistance. Here, transcriptomic responses of the two fungal pathogens Blumeria graminis f. sp. hordei (barley powdery mildew) and Puccinia triticina (wheat leaf rust) during growth on their respective host plants containing Lr34 were compared to their responses on control plants without Lr34. Two different time points after inoculation were chosen for analysis of powdery mildew on barley and one time point for wheat leaf rust. Transcriptome analyses revealed that there were no differences in the expression patterns of the two pathogens growing on susceptible versus partially resistant plants, even though pathogen growth was reduced in the presence of Lr34. This reflects the absence of observable reaction in the pathogen to the presence of the Lr34 resistance gene and, consequently, no major alteration of fungal pathogen metabolism.

Downy mildew, commercially the most important disease of spinach, is caused by the obligate oomycete Peronospora effusa. In the past two decades, new pathogen races have repeatedly overcome the resistance used in newly released cultivars, urging the need for more durable resistance. Commercial spinach cultivars are bred with major R genes to impart resistance to downy mildew pathogens and are effective against some pathogen races/isolates. This work aimed to evaluate the worldwide USDA spinach germplasm collections and commercial cultivars for resistance to downy mildew pathogen in the field condition under natural inoculum pressure and conduct genome wide association analysis (GWAS) to identify resistance-associated genomic regions (alleles). Another objective was to evaluate the prediction accuracy (PA) using several genomic prediction (GP) methods to assess the potential implementation of genomic selection (GS) to improve spinach breeding for resistance to downy mildew pathogen. More than four hundred diverse spinach genotypes comprising USDA germplasm accessions and commercial cultivars were evaluated for resistance to downy mildew pathogen between 2017–2019 in Salinas Valley, California and Yuma, Arizona. GWAS was performed using single nucleotide polymorphism (SNP) markers identified via whole genome resequencing (WGR) in GAPIT and TASSEL programs; detected 14, 12, 5, and 10 significantly associated SNP markers with the resistance from four tested environments, respectively; and the QTL alleles were detected at the previously reported region of chromosome 3 in three of the four experiments. In parallel, PA was assessed using six GP models and seven unique marker datasets for field resistance to downy mildew pathogen across four tested environments. The results suggest the suitability of GS to improve field resistance to downy mildew pathogen. The QTL, SNP markers, and PA estimates provide new information in spinach breeding to select resistant plants and breeding lines through marker-assisted selection (MAS) and GS, eventually helping to accumulate beneficial alleles for durable disease resistance.

An F4-derived F6 recombinant inbred line population (n = 148) of a cross between the durable stripe (yellow) rust (caused by Puccinia striiformis) and leaf (brown) rust (caused by Puccinia triticina) resistant cultivar, Triticum aestivum 'Cook', and susceptible genotype Avocet-YrA was phenotyped at several locations in Canada and Mexico under artificial epidemics of leaf or stripe rusts and genotyped using amplified fragment length polymorphism (AFLP) and microsatellite markers. Durable adult plant resistance to stripe and leaf rusts in 'Cook' is inherited quantitatively and was based on the additive interaction of linked and (or) pleiotropic slow-rusting genes Lr34 and Yr18 and the temperature-sensitive stripe rust resistance gene, YrCK, with additional genetic factors. Identified QTLs accounted for 18% to 31% of the phenotypic variation in leaf and stripe rust reactions, respectively. In accordance with the high phenotypic associations between leaf and stripe rust resistance, some of the identified QTLs appeared to be linked and (or) pleiotropic for both rusts across tests. Although a QTL was identified on chromosome 7D with significant effects on both rusts at some testing locations, it was not possible to refine the location of Lr34 or Yr18 because of the scarcity of markers in this region. The temperature-sensitive stripe rust resistance response, conditioned by the YrCK gene, significantly contributed to overall resistance to both rusts, indicating that this gene also had pleiotropic effects.

Stripe rust of wheat, caused by Puccinia striiformis f. sp. tritici, continues to cause severe damage worldwide. Durable resistance is necessary for sustainable control of the disease. High-temperature adult-plant (HTAP) resistance, which expresses when the weather becomes warm and plants grow older, has been demonstrated to be durable. We conducted numerous studies to understand the molecular mechanisms of different types of stripe rust resistance using a transcriptomics approach. Through comparing gene expression patterns with race-specific, all-stage resistance controlled by various genes, we found that a greater diversity of genes is involved in HTAP resistance than in all-stage resistance. The genes involved in HTAP resistance are induced more slowly and their expression induction is less dramatic than genes involved in all-stage resistance. The high diversity of genes and less dramatic induction may explain durability and the incomplete expression level of HTAP resistance. Identification of transcripts may be helpful in identifying resistance controlled by different genes and in selecting better combinations of genes to combine for achieving adequate and durable resistance.

Chapter 11 - Wheat rust research: impact, thrusts, and roadmap to sustained wheat production

Changes in the genetic structure of the lettuce crop ( Lactuca sativa ) and populations of the lettuce downy mildew pathogen ( Bremia lactucae ) in Germany from 1974 to 1997 were studied and the genetic and population genetic results of race surveys of B. lactucae were reinterpreted. Data on 176 B. lactucae isolates collected from L. sativa cultivars at 48 sites in 11 German federal states were analysed. The host population contained 12 well‐defined race‐specific resistance genes ( Dm 1, Dm 2, Dm 3, Dm 4, Dm 5/8, Dm 6, Dm 7, Dm 11, R12, Dm 13, Dm 16 and R18). However, only some of these genes (e.g. Dm 3, Dm 7, Dm 11, Dm 16 and R18) played an important role in management of resistance in the host crop. Substantial changes in the frequencies of these R‐genes were recorded over the period as a reaction to the occurrence of new virulent phenotypes in the pathogen population. Generally, the durability of resistance was very short. However, distinct regional patterns were observed for the frequencies of different virulence (v) factors. The data on changes in the virulence of B. lactucae population exemplified well the process of host–pathogen coevolution. During the period studied, there was a considerable increase in the frequency of some v‐factors (v2, v3, v5/8, v6, v7, v11 and v16) and in the complexity of v‐phenotypes, while the diversity in the pathogen population in general decreased. In 1996/97, 12 of the 14 v‐factors evaluated reached frequencies higher than 0·90 in the pathogen population. Relatively low frequencies were observed only for factors v14 (0·17) and v18 (0·24). The presence of 23 v‐factors was studied in six German isolates virulent on lettuce cultivar Titan, possessing the resistance gene R36 derived from Lactuca saligna and introduced in German lettuce crops during the mid‐1990s. Isolate DEG2 showed the highest complexity of v‐phenotype; it overcame the resistance in cv. Mariska (R18) and line CS‐RL (R18+?), but some sporulation occurred also on cv. Titan ( Dm 6 + R36). Conversely, the isolates SR2 and SAW1 overcame resistance based on R36, but were avirulent to R18. Future utilization of R‐genes in commercial lettuce cultivars is proposed; R‐genes R18 and R36 could be considered most efficient for this purpose. However, they may be effective only in certain regions.

Three rusts are destructive, diminishing produce and nutritious value significantly, affect food availability and consequently food security through reductions in yield. In agricultural research institutes with mandate of wheat improvement, incorporating genes resistant against rust is matter of routine. The dilemma of rusts in wheat has been addressed the most, leading to discovery of principles of plant breeding for resistance e.g. gene disease genes inherit following Mendelian genetics, concept of genetic diversity and concept of gene for gene theory. Two strategies of breeding wheat for disease resistance are being followed. 1- Conventional and 2- Advanced. Among conventional approaches selection and hybridization are well known. However rust resistance has been found short lived and may also be durable in certain cases. Durability of disease resistance is desired and has been explored widely. Durability of resistance is generally attained through incorporation of genes effective at adult plant stage and combination of quantitative genes. Application of biotechnology to improve productivity of rust resistance breeding is the usage of molecular markers in pyramiding genes and substantiates the existence of genes in, and confirming released cultivars are pure. This involves molecular markers that are precise and pertinent across extensive ranges of breeders’ germplasm. This review article encompasses all features of wheat development through application of different techniques of wheat improvement. However, despite development of novel approaches that has accelerated wheat breeding, breeding in pathogen leading to producing more virulent strains. Consequently, wheat breeding is a continuous process.

Both hypersensitive and quantitative forms of resistance to the bacterial spot pathogens (Xanthomonas spp.) occur in pepper and tomato. Five resistance genes involved in hypersensitivity in pepper and four in tomato have been identified so far. The corresponding pathogen avirulence genes have been cloned and characterized, and features, including a propensity for accumulating mutations and at times, loss of plasmid-borne avirulence genes, are known to occur. The frequency of these changes affects race composition among pathogen populations and determines the durability of the corresponding plant resistance. At least four different species of Xanthomonas are known to cause bacterial spot, and these can differ in specific avirulence gene content. Quantitative or multigenic resistance has also more recently been researched and appears to be more durable than the hypersensitive resistance. Two recessive genes have been identified that yield a nonhypersensitive form of resistance in pepper and together can provide strong resistance. More emphasis is being given to transfer of quantitative trait resistance to commercial cultivars of both tomato and pepper.

Yellow rust, caused by Puccinia striiformis, is one of the most damaging diseases affecting bread wheat in temperate regions. Although resistance to yellow rust is frequently overcome by new virulent races, a durable form of resistance in the French bread wheat Camp Remy (CR) has remained effective since its introduction in 1980. We used 217 F7 recombinant inbred lines (RILs) derived from the cross between CR and the susceptible cultivar Recital to identify and map quantitative trait loci (QTLs) involved in durable yellow rust resistance. Six significant QTLs that were stable over a 4-year period were detected. Two QTLs, denoted QYr.inra-2DS and QYr.inra-5BL.2, were located on the short arm of chromosome 2D and the long arm of chromosome 5B, respectively. Each explained on average 25-35% of the observed phenotypic variation and were probably inherited from Cappelle Desprez, a parent of CR that confers durable adult plant resistance to yellow rust. QYr.inra-2DS probably corresponds to the Yr16 gene. The most consistent QTL, designated QYr.inra-2BL, was located on the centromeric region of chromosome 2B and explained 61% of the phenotypic variation in 2003. This QTL was responsible for seedling-stage resistance and may correspond to a cluster of genes, including Yr7. The remaining QTLs were mapped to the short arm of chromosome 2B (R2=22-70%) and to the long arm of chromosomes 2A (R2=0.20-0.40) and 5B (R2=0.18-0.26). This specific combination of seedling and adult plant resistance genes found in CR and CD may constitute the key to their durable resistance against yellow rust.

THE TITLE of this paper sounds as if the roof were about to fall in on us-that we have lost or are about to lose the fight with the fungi. Say not so! We have only begun to fight, but fight we mnust. We may easily forget that fungi feed at the same table with us. This is so because they have a ticket to the first sitting. They consume our food in the farmer's field, on the trains and trucks that bring it to us, and in the grocer's store. If this fight go not forward to success, we may one day not be able to smile at the naivete of Mr. Malthus who thought that we would soon eat ourselves out of our own food supply. Fungi have been on this planet longer than we. They have developedsome fantastically efficient devices that serve them in their fight with us. They are well able to search out our food plants so that they also may eat, drink and be merry. It has been fun to help a little in the research to develop the counter measures that we use in our fight with them. Before we come to the counter measures, however, I should like to discuss some of the famous plant diseases of antiquity and how some of them have altered the course of history. Three plants diseases of modern times are known to almost everyone. Perhaps the best known isthe chestnut blight that swept every chestnut tree from the hills from Maine to Georgia. The second is the Dutch elm disease that is marching down the streets of cities and killing the elms from Montreal to Denver. And the third is oak wilt. It is scaring the wits out of the people who produce the oak flooring for our houses and the kegs for our beer. These diseases latch onto our consciousness because they are new and they strike down handsome big trees. These are some of the,blasts and blights that beset us, but what really robs us are such diseases as wheat rust and potato rot. Wheat rust is perhaps the most famous disease of antiquity and it is still with us. Wheat rust robs us of our bread, the very staff of life. Those of us who went through both World Wars remember the of World War I. We had wheatless days in that war because 1917 was one of those years when wheat rust swept the plains and consumed the grains like a prairie fire. The wheat rust fungus ate most of our bread at the first sitting. We had to settle for rice and corn bread. We were lucky during World War II. Wheat rust did not stage another such ruinous raid and we did not have wheatless days. Of course, scientific research had also been at work in the meantime and had won part of the fight with that fungus. Wheat rust was known to the Israelites who talked about it in Genesis. It was known to the Romans. And it was known in Colonial America. On this last point, there hangs a tale of the impact of a plant disease on civilization. This is the tale of how wheat rust altered the eating habits of a group of people. When the English colonists came to America, some settled in New England, and others in tidewater Virginia, Plymouth and Jamestown being settled within a few years of each other. Undoubtedly, both groups of immigrants brought wheat with them, and they both found the Indians growing corn. The wheat rust disease, however, acted differently on the wheats in the two colonies, just as it does today.

ABSTRACTLeaf rust and powdery mildew, caused by Puccinia triticina and Blumeria graminis f. sp. tritici, respectively, are widespread fungal diseases of wheat (Triticum aestivum L.). Development of cultivars with durable resistance is crucially important for global wheat production. This paper reviews the progress of genetic study and application of adult plant resistance (APR) to wheat leaf rust and powdery mildew. Eighty leaf rust and 119 powdery mildew APR quantitative trait loci (QTL) have been reported on 16 and 21 chromosomes, respectively, in over 50 publications during the last 15 yr. More important, we found 11 loci located on chromosomes 1BS, 1BL, 2AL, 2BS (2), 2DL, 4DL, 5BL, 6AL, 7BL, and 7DS showing pleiotropic effects on resistance to leaf rust, stripe rust, and powdery mildew. Among these, QTL on chromosomes 1BL, 4DL, and 7DS also correlate with leaf tip necrosis. Fine mapping and cloning of these QTL will be achieved with the advent of cheaper high‐throughput genotyping technologies. Germplasm carrying these potential resistance genes will be useful for developing cultivars with durable multidisease resistance. In addition to its non‐NBS–LRR (nucleotide binding site–leucine rich repeat) structure, the senescence‐like processes induced by Lr34 could be the reason for durability of resistance; however, more information is needed for a full understanding of the molecular mechanism related to durability. Adult plant resistance genes have been used by CIMMYT for more than 30 yr and have also been transferred to many Chinese wheat varieties through shuttle breeding.

Background: At all stages of their development, plants are in permanent contact with causative agents of various diseases. Mechanisms of disease resistance and its durability in crops largely depend on the pathogen’s lifestyle, namely the nutrition mode and host range. Objective: The objective of this review is to consider the main advances in the production of genotypes with durable disease resistance in the globally important food crops, wheat, rice, and potato, as well as barley. Results: In wheat, durable resistance could be provided by the employment of various adult plant resistance genes against biotrophic pathogens, whose action commonly does not involve hypersensitivity response, as well as major quantitative genes, including mutants of susceptibility alleles, against necrotrophs via marker assisted selection (MAS). In barley, the most prominent example is the gene mlo conferring durable powdery mildew resistance, but it is compromised by higher susceptibility to some necrotrophic fungi. A few genes for broad-spectrum resistance against the rice blast and bacterial blight pathogens confirmed their effectiveness for decades, and they could be combined with effective R genes via MAS. Resistance to late blight of potato is mainly provided by R genes introgressed from wild potato species, which could be pyramided with quantitative trait loci. Genes for extreme resistance to potato viruses derived from related species provide durable and broad-spectrum resistance and could be effectively deployed in potato breeding using MAS. Silencing susceptibility genes by genome editing technologies is the most promising approach to produce plants with durable resistance to many pathogens in the crop species. Genetic transformation with genes for resistance-associated proteins or constructs providing silencing via RNA interference is an effective biotechnological method to generate plants with durable resistance against pathogens, especially viruses. Conclusion: Main advances in the production of crop plants with durable resistance are based on studies of molecular mechanisms of plant immunity and its special features for pathogens with different lifestyles via the use of biotechnological approaches such as MAS for pyramiding of monogenic quantitative resistance genes or qualitative R genes, changes in expression of certain genes associated with resistance, the introduction of transgenes, mutagenesis and genome editing aimed at silencing susceptibility genes.