Major Genes for Powdery Mildew Resistance in Research and Breeding of Barley: A Few Brief Narratives and Recommendations

1. Introduction

To breed varieties resistant to harmful diseases is an important aim in crop breeding. However, pathogens have different life cycles and adaptation mechanisms (evolutionary forces) which are important in the promotion of population diversity and adaptability [1,2,3,4]. These aspects explain why strategies of crop protection, including the use genetic resistance in different plant pathosystems, can differ.

In the Czech Republic (central Europe) an extensive study and practical use of major resistance genes in barley (Hordeum vulgare L.) against powdery mildew caused by Blumeria hordei (M. Liu & Hambl.) has resulted in increased knowledge, experiences and conclusions [5,6,7,8,9,10,11,12,13,14] that are explored in this review.

Despite the drawbacks of major genes of specific resistance against barley powdery mildew described below, some research laboratories and breeding companies are still using these resistances for varietal production. This report will outline why this strategy should be modified in favor of adopting non-specific and durable resistance.

Co-evolution of barleys carrying specific resistances and local powdery mildew populations has created wide pathogen variation. Here is also shown that this reflection major genes of specific resistance in the pathogen population can be utilized for postulating (identifying) major resistance genes and their combinations and a knowledge of the host variation can assist in the uncovering of varietal non-authenticity and genotype heterogeneity including gene bank accessions. Hence, pathotypes selected from populations of the pathogen which caused “bust” (overcome) major genes of specific resistance and resulted in losses on barley production can be a tool for resistance gene postulation and thus prevent losses through the use of non-authentic genotypes in research and breeding.

This paper has three closely linked aims, i) to present convincing examples of the rapid breakdown of race-specific resistances due to great adaptability of the pathogen, which lead to financial losses of growers, seed companies and breeders, ii) to show usefulness of using pathotypes of the pathogen that mostly developed from the breakdown of specific resistances to postulate major genes which can avert financial losses in barley research and breeding, and iii) to recommend the adoption of durable resistances against powdery mildew for barley improvement.

2. Major Genes and Crop Resistance

Many major genes of crop resistances to diseases have been detected. A common feature of genes selected for breeding resistant varieties is their efficiency against tested pathogen populations. Such genes provide resistance to varieties in the field, but usually for a limited time.

Resistance genes are denoted “major” because of their great effectiveness against disease. Major genes of specific resistance have two dimensions, namely resistance to a disease when infected by a limited number of pathotypes or by pathogen populations.

2.1. Non-Durable Major Resistance Genes

An example of the highest efficiency of a barley major resistance gene against powdery mildew is the allele Mla8 characterized by the lowest phenotype – infection response 0 (IR0), which is manifested by no traces of the disease after inoculation of host genotypes with avirulent pathotypes (Figure 1). However, isolates avirulent to Mla8 are extremely rare (see later) and are absent in the field. Therefore, this gene is ineffective and varieties containing Mla8 are fully susceptible to “natural” pathogen populations worldwide.

There are numerous well-characterised hypersensitive major resistance genes against barley powdery mildew [15,16] and many others are present in wild barley (H. vulgare subsp. spontaneum) [17,18,19]. However, they conform to the principles of the gene-for-gene system whereby each resistance gene of the host corresponds with a virulence gene of the pathogen [20] and their resistance is race-specific (denoted “specific”). When used in commercial varieties their initial resistance is soon overcome as a result of an increasing frequency of virulent pathotypes which are under strong directional selection once varieties carrying such genes are grown [21]. The following examples illustrate the outcomes and conclusions drawn from cultivation of varieties with major resistance genes.

In 1965 a domestic spring barley variety Diamant – an X-ray mutant from an old variety Valticky was registered. It possessed a new semi-dwarf sdw1.d allele [22]; varieties bred from it are characterized by “Diamant (morpho)type” and these semi-dwarf derivatives are often called “Diamant series” (of varieties). Diamant has made a positive contribution to crop breeding worldwide, but is susceptible to powdery mildew. This was the commonest disease of cultivated barley in Europe, especially in central and northwest regions [2,23] until the expansion of spring varieties carrying a nonspecific resistance. Diamant contains the ineffective major resistance gene Mla8 and its average scoring in 161 field trials conducted in 1971–1975 was 3.77 on a nine point scale where 9 is the highest resistance [24]. The high susceptibility of Diamant encouraged breeders to combine desirable agronomical characteristics of this variety with effective resistance against the disease.

In 1977 Spartan – a descendant of Diamant was released that carried a “new” gene Mla9 and also Mlk1 [13] providing an overall resistance of 8.60 in 39 field trials conducted a year before its registration [24]. Three years later Spartan was historically the most widely-grown variety in the Czech Republic occupying 191.000 ha (31.9% of the crop area). The following year, when its area was 190.000 ha, the resistance was overcome (an average field scoring of 4.53) and in 1983 Spartan became the most susceptible (3.38) among all tested varieties. Since Spartan was the only commercial variety possessing Mla9 and because other resistant varieties were already available (see next paragraph) its growing area decreased quickly to 1.2% in 1985. The widespread cultivation of Spartan and progression of its susceptibility demonstrate a typical ‘boom-and-bust’ cycle of varieties with specific major resistance genes against diseases caused by pathogens that rapidly adapt [3].

Another example is Koral released in 1978, in which, similarly to Spartan, Mla8 of its ancestors was substituted with another allele, in this case Mla13 and simultaneously carrying Mlg [13]. Koral was the first central European variety possessing Mla13 [25] and its resistance averaged 8.93 in 105 field trials carried out in 1976–1978 [24]. Until 1985 five varieties having Mla13 were registered in the Czech Republic out of a total of 13 varieties registered in this eight year period [13]. Varieties with Mla13 were the most resistant for a decade (1976–1985) [24] and up to 1985 they were grown on a total of about 1.5 Mha. After some infection in 1985 their resistance fully broke down in 1986 when these varieties occupied the highest proportion (63.5%) of the total crop area 445 000 ha [9]. The years 1987–1989 had the 4th, 1st and 2nd strongest mildew epidemics, respectively, that were recorded on spring barley over a thirty year period (1976–2005) [23], and in 1989–2000 varieties carrying Mla13 were the most susceptible in eight of these 12 years [24]. Although the resistance was already ineffective, susceptible varieties possessing Mla13 were still grown and occupied 61.0% of the crop area in 1990 [9]. In addition more varieties with Mla13 were registered (10 until 1996 and three more up to 2011) because until 1993 there were no other commercial resistant varieties available [13,26]. The total area of Mla13 varieties grown in 1986–2005, when they were fully susceptible, was 2.6 Mha, thus much larger than the area when they were resistant. The rapid spread of airborne spores of pathotypes containing important virulences (Va13 and Vg as well as Va6, Va7, Va9 and Vk1) produced on these widely-grown and already fully susceptible varieties caused strong powdery mildew epidemics in large parts of Europe in the second half of 1980ʼs [27].

In the domestic population virulence against a major resistance gene Mlp was not found before 2012, but soon after the introduction of winter barley varieties carrying this gene the frequency of corresponding virulence reached almost 70% [12]. The fate of varieties possessing other major genes of specific resistance, even those grown on small areas, was similar [24]. It raises questions about the benefit of major genes of specific resistance against barley powdery mildew.

Despite these examples and many other instances of specific resistances of barley rapidly losing their control of the disease, there are still research programs using this outdated strategy [28,29,30,31,32,33].

2.2. Durable Major Resistance Genes

Only one durable, non-specific and non-hypersensitive major resistance of barley against powdery mildew that does not conform to the gene-for-gene concept is known [34,35]. It is monogenic, conditioned by one of many recessive genes of similar function [15] and designated Mlo [36] (functional genes are designated mlo). In commercial varieties mlo11 derived from Ethiopian landraces was the first to be exploited and subsequently mlo9 from a mutant SZ 5139 derived from Diamant was adopted by breeders who released Alexis. This was the most widely grown European variety in the latter part of the 1980s [35,37]. Conversely, from the same set of mutants a powdery mildew super-susceptible line SM 4142 was selected and has been used in our laboratory.

Since 1993 164 spring barley varieties bred in nine central and northwest European countries have been registered in the Czech Republic, 114 of them (69.5%) carrying Mlo resistance. During 2021–2023 almost 96% possessed Mlo and in registration trials such varieties have been the most resistant since 1985 [21]. Despite its long-term durability there is some experimental evidence that the pathogen can develop pathotypes partially virulent to Mlo [38,39,40,41]. There could be a higher risk for this to occur in areas where spring and winter barleys with this resistance are concurrently grown.

Jørgensen [35] predicted that Mlo resistance should also be present in other plants and this was confirmed when it was documented in some species in addition to barley [42]. With continuing research the number of plant species with detected Mlo resistance is increasing [43,44,45,46,47,48,49,50,51,52].

2.3. Pathogen Variation

Successful postulation (see section 2.4) of major resistance genes in varieties depends on the use of wide virulence variation of the pathogen as outlined in the following examples.

The first European commercial varieties carrying introduced resistance genes were registered in the 1950s since prior to this date no specific varietal resistance was detected However, isolates avirulent on many varieties including some of European origin were found in Japan and these isolates enabled the discovery of the gene Mla8 [53]. In Denmark, a set of European varieties was tested with one of these Japanese isolates (Race I) and from the results it was concluded that Mla8 is frequently present in old spring barley varieties [54]. In varieties bred subsequently Mla8 was almost always replaced with effective Mla alleles at the same locus [25] as in the cases of Spartan and Koral mentioned above.

Another example relates to discussions held at the International Center for Agriculture Research in the Dry Areas (ICARDA) in 2004. Tadmor, a susceptible barley variety selected from a Syrian landrace Arabi Aswad [S. Ceccarelli, personal communication], was earmarked as a suitable host for powdery mildew experimental research. In resistance tests with about 50 isolates Tadmor was fully susceptible (IR4) to all isolates except Race I, to which it was fully resistant (IR0) and had an identical infection response array (IRA) to varieties possessing Mla8. However, in subsequent tests a few Israeli isolates were included and the set of barleys with IRAs previously indicating the presence of Mla8 was split into two groups. Varieties in the first group had a similar IRA as before when only Race I was avirulent (Mla8), whereas varieties in the other group, including Tadmor, were fully resistant (IR0) to this isolate as well as to two Israeli isolates. Thus, the gene present in the second group differed from Mla8 and this newly-discovered resistance was tentatively named after Tadmor (Ta). Despite this, a commercial winter barley variety Lomerit was present in the same group and it was considered more appropriate to designate the gene accordingly (MlLo). Subsequent research revealed that MlLo is an allele or pseudo-allele of the Mla locus [11,16] and although MlaLo was undetected until recently, tests of gene bank accessions showed that Lo is the most frequent major resistance present alone or in combinations with other powdery mildew resistances in more than 26% genotypes derived from old and current winter barley varieties [Dreiseitl and Nesvadba, unpublished].

Recent research confirmed that the central European barley powdery mildew population contains the widest spectrum of known virulences and their combinations among all regions where barley is grown and that it is one of most diverse populations among plant pathogens [12]. Selected isolates from this population are important for postulating the composition of complex resistance gene combinations. On the other hand, the most frequent genes in spring and winter barley (Mla8 and MlaLo) could be revealed only by using non-European isolates because these avirulences are absent in Europe.

2.4. Postulation of Major Resistance Genes

Classical identification of major resistance genes in host varieties is based on the phenotypic responses of varieties after inoculation with pathogen isolates to obtain a row of IRs for each genotype. Comparing IRAs of tested varieties with IRAs of standard lines possessing known resistance genes can identify genes and their combinations [6,13,30,55,56,57,58] including designating new resistances [59]. This method denoted as the “postulation” of major resistance genes [60] is widely used to characterize genes in cereals against biotrophic pathogens such as powdery mildews and rusts [61,62,63,64,65,66,67]. In the case of mildews the term “postulation” was introduced later, first for mildew on wheat [68,69,70] and subsequently for barley mildew [10,71,72].

An integral part of exploiting major resistance genes is their detection. Molecular markers [73,74,75,76,77] are helpful tools mainly as an aid to select lines with the required gene(s). However, to distinguish new and more complex gene combinations postulation is still the most suitable way, especially if maximum pathogen variation is used. Gene postulation based on a gene-for-gene concept in association with Mendelian genetic analysis was also used to clarify complex resistances in some accessions including highly heterogeneous ones [11]. The host powdery mildew resistance variation [15] was recently updated [16].

2.5. Other uses of Major Resistance Genes

Plant research and breeding depends on accessing a range of diverse plant genotypes that are available in gene banks. Non-authenticity of gene bank accessions due to human error is one of the most serious problems [10,78,79,80,81,82,83,84]. To overcome this shortcoming there are modern efficient methods for verifying the identity of plant varieties including sequencing or protein spectra analyses [85,86]. However, these methods are often unsuitable for older accessions lodged in gene banks as they may be incorrectly labelled or contaminated with other genotypes [87] because of insufficient historical data to compare with results obtained with modern techniques. On the other hand, there is a lot of information about the presence of major resistance genes against powdery mildew in barleys when varieties were registered or collected and for almost 700 European varieties such data were summarized [25]. Therefore, results of current postulation of major resistance genes in accessions, including those from gene banks, can be compared with original data and on this basis varietal authenticity established. The problem is well illustrated in the following three examples resulting from the study of spring barley gene bank accessions.

For resistance gene postulation of accessions maintained in a gene bank five SSPs of each accession were tested. The set of varieties included Abyssinian 1102, an Ethiopian landrace containing mlo11, one of two most important recessive resistance genes against barley powdery mildew and one of numerous sources of Mlo [35]. Three different genotypes were found among its single seed progenies (SSPs) but none of them carried mlo [88]. This proves that this accession was heterogeneous and incorrectly labelled “Abyssinian 1102”.

Second, an accession of Diamant was studied [88]. Three lines (SSPs) were eliminated since they contained mlo and Mla7, both used in breeding barley varieties subsequent to Diamant [25]. Two SSPs carried Mla8 and only these can be considered as the true variety.

The last example is from a recent study comparing accessions from the domestic gene bank with those from foreign gene banks [87]. Asse (HOR 4482), is almost six decades old German variety that contains Mlg [89]. In our studies we found Mla8 in the USA accession and Mlra in the accession from the Czech gene bank. Because no true genotype was present in Asse accessions, a sample of this variety should be obtained from other gene banks, preferably from its country of origin.

These examples clearly demonstrate that a knowledge of major resistance genes present in varieties is crucial. Even those genes whose resistance has broken down and are already ineffective are useful as “markers” or “information tools” for identifying varieties and confirming their pedigree, genotype homogeneity and authenticity.

3. Conclusions

Breeding barley against powdery mildew, especially in Europe, has been based almost exclusively on exploiting major resistance genes. Unlike the later use of non-specific resistance Mlo, genes of specific resistance bring a short-term protective effect terminated by the breakdown of resistance in the field and consequently associated with risk and financial losses for growers and breeding companies.

Other significant errors occur when mislabeled or genetically heterogeneous varieties confound research aims or breeding projects and lead to false scientific results and conclusions. This review presents examples of the first case (‘boom-and-bust’ cycle of varieties with major genes of specific resistance) and summarizes the root causes and dangers using of non-authentic varieties which can be detected by identifying major resistance genes against powdery mildew and highlights ways to avoid associated losses.

4. Future Directions

Most of the known major resistance genes of barley against powdery mildew are race-specific and short-lived. The only exception is a major non-specific resistance Mlo, which has remained durable after almost a half of century of its widespread use mainly in central and western Europe where concurrently grown spring and winter barley is commonly practised and conditions for the pathogen are favorable. In such an environment consisting of an uninterrupted green bridge it is recommended that Mlo should only be present in spring varieties to prevent a possible slow partial adaptation of the pathogen. For winter barley it is advisable to accumulate quantitative (minor) non-specific resistance genes [90,91], or develop effective resistances introgressed from Hordeum bulbosum [92,93,94,95] or, if it were possible, from species in the tertiary genepool of barley that are immune to closely related pathogens.