Mapping quantitative trait loci associated with spot blotch and net blotch resistance in a doubled-haploid barley population

Spot blotch and net blotch are important foliar barley (Hordeum vulgare L.) diseases in Canada and elsewhere. These diseases result in significant yield reduction and, more importantly, loss of grain quality, downgrading barley from malt to feed. Combining resistance to these diseases is a breeding priority but is a significant challenge using conventional breeding methodology. In the present investigation, an evaluation of the inheritance of resistance to spot and net blotch was conducted in a doubled-haploid barley population from the cross CDC Bold (susceptible)?×?TR251 (resistant). The population was screened at the seedling stage in the Phytotron and at the adult-plant stage in the field for several years. Chi-squared analysis indicated one- to four-gene segregation depending on disease, isolate, plant development stage, location and year. A major seedling and adult-plant resistance quantitative trait locus (QTL), designated QRpt6, was re-confirmed for net-form net blotch resistance, explaining 32?C61% of phenotypic variation in different experiments. Additional QTL for seedling and adult-plant resistance to net blotch were identified. For spot blotch resistance, a major seedling resistance QTL (QRcss1) was detected on chromosome 1H for isolate WRS1909, explaining 79% of the phenotypic variation. A highly significant QTL on 3H (QRcs3) was identified for seedling resistance to isolate WRS1908 and adult-plant resistance at Brandon, MB, Canada in 2008. The identification of QTL at only one location or from 1?year suggests spot blotch resistance is complex and highly influenced by the environment. Efforts are being made to combine spot and net blotch resistance in elite barley lines using molecular marker-assisted selection.     

Ultrastructural and Immunocytochemical Studies on Effects of Barley Yellow Dwarf Virus – Infection on Fusarium Head Blight, Caused by Fusarium graminearum, in Wheat Plants

The interactions between barley yellow dwarf virus (BYDV) and Fusarium head blight (FHB), caused by Fusarium graminearum, were studied in the two winter wheat cultivars (cvs.), Agent (susceptible to FHB) and Petrus (moderately resistant to FHB), using ultrastructural and immunocytochemical methods. Infections of wheat plants of both cvs. by BYDV increased susceptibility to FHB. BYDV infection caused numerous cytological changes in lemma tissue of both cvs. such as formation of vesicles in the cytoplasm, degradation of fine structures of chloroplasts of both cvs. and accumulation of large starch grains in the chloroplasts. Electron microscopical studies showed that the development of F. graminearum on spike surfaces was not affected in BYDV‐infected plants. After penetration and intercellular growth in lemma tissue, defence responses to Fusarium infections were markedly reduced in BYDV‐diseased plants compared to the tissue of virus‐free plants. At sites of contact of fungal cells with host tissue, depositions of cell wall material were distinctly less pronounced than in tissues of virus‐free plants of cv. Petrus. Detection of β‐1,3‐glucanases and chitinases in lemma tissue of cv. Agent revealed no appreciably increased accumulation of both defence enzymes in F. graminearum‐infected virus‐free and BYDV‐infected tissues compared to the non‐infected control tissue. On the other hand, in cv. Petrus, infection with F. graminearum induced a markedly enhanced activity of both enzymes 3 days after inoculation. The increase of both enzyme activities was less pronounced in BYDV‐infected plants than in tissue exclusively infected with F. graminearum. Cytological studies suggest that in contrast to the susceptible cv. Agent postinfectional defence responses may play still an important role in the resistance of the moderately resistant cv. Petrus to FHB.     

A biological and molecular characterization of some Egyptian barley genotypes which are resistant to net blotch disease

A survey for resistance against net blotch disease (caused by Pyrenophora teres) was performed on some Egyptian barley landraces and some selected resistance and susceptible standard German barley genotypes. The results indicated that most of the Egyptian barley landraces are extremely resistant to the disease. Molecular analysis using RAPD and AFLP showed unique banding profiles for the different genotypes, and specific AFLP markers for the Egyptian genotypes were identified. The effectiveness of RAPD and AFLP for identifying different barley genotypes of different origins and with different reactions against P. teres was discussed. The results of the biological evaluation and molecular characterization done in this study can be seen as the starting point needed to identify the valuable net blotch resistant Egyptian barley germplasm at both the phenotype and genotype levels and draw the attention of breeders and banks of natural plant genetic resources towards this valuable yet neglected germplasm.     

A novel cost of R gene resistance in the presence of disease

Resistance responses can impose fitness costs when pests are absent. Here, we test whether the induction of resistance can decrease fitness even in plants under attack; we call this potential outcome a net cost with attack. Using lines in which genetic background was controlled, we investigated whether susceptible Arabidopsis thaliana plants can outperform R gene resistant plants when infected with pathogens. For the R gene RPS2, there was a fitness benefit of resistance in the presence of intraspecific competition, but there was a net cost in the absence of competition: resistant plants produced less seed than susceptible plants even though infected with Pseudomonas syringae. This net cost was primarily due to overcompensation by susceptible plants, which occurred because of a developmental response to infection. For the R gene RPP5, there was no fitness effect of resistance without competition but a net cost when plants were infected with Peronospora parasitica in the presence of competition. This net cost was due to a reduction in the fitness of infected, resistant plants and complete compensation in susceptible plants. A spatially variable model suggests that a trade-off between net benefits and net costs with attack may help explain the persistence of individuals lacking R gene resistance to disease.     

Interaction between Rhynchosporium secalis and Pyrenophora teres in the Field and Identification of Genotypes with Double Resistance in a Doubled-haploid Barley Population

Net blotch [Pyrenophora teres (Died.) Drechsl.] and scald [Rhynchosporium secalis (Oudem.) J.J. Davis] are the two most important foliar diseases of barley (Hordeum vulgare L.) in Tunisia. The use of cultivars with double resistance is the most effective method in controlling both diseases. A doubled‐haploid barley population derived from Tunisian cultivars was evaluated to both net blotch and scald during two growing seasons in the field. Mass disease index (MDI), area under the disease progress curve (AUDPC) and apparent infection type (r) were used to assess disease reaction. MDI of net blotch and scald reached up to 65% and 90% respectively. Least significant difference (LSD) test and comparison of the reaction of the doubled haploid (DH) lines to the overall population mean value were efficient in identifying lines with double resistance to both diseases. From the 59 DH lines screened, lines 21, 33, 37, 46 and 47 showed the best level of adult plant resistance to both diseases and may be used in a breeding program for diseases resistance. Interactions between R. secalis and P. teres were investigated at the level of the whole plant under variable epidemic conditions. Under low epidemic conditions, net blotch and scald developments were usually independent, but positively associated for tolerant lines for both diseases. Under high epidemic conditions, competition effects were obtained for susceptible and resistant genotypes. This competition seems to be an exploitation competition that is associated with decreasing resource availability as it occurs only with high levels of infestation or/and when susceptible lines are infected. This study demonstrates the variability of net blotch and scald interaction with epiphytotic conditions and group of resistance.     

Mapping quantitative trait loci associated with barley net blotch resistance

Net blotch of barley, caused by Pyrenophora teres Drechs., is an important foliar disease worldwide. Deployment of resistant cultivars is the most economic and eco-friendly control method. This report describes mapping of quantitative trait loci (QTL) associated with net blotch resistance in a doubled-haploid (DH) barley population using diversity arrays technology (DArT) markers. One hundred and fifty DH lines from the cross CDC Dolly (susceptible)/TR251 (resistant) were screened as seedlings in controlled environments with net-form net blotch (NFNB) isolates WRS858 and WRS1607 and spot-form net blotch (SFNB) isolate WRS857. The population was also screened at the adult-plant stage for NFNB resistance in the field in 2005 and 2006. A high-density genetic linkage map of 90 DH lines was constructed using 457 DArT and 11 SSR markers. A major NFNB seedling resistance QTL, designated QRpt6, was mapped to chromosome 6H for isolates WRS858 and WRS1607. QRpt6 was associated with adult-plant resistance in the 2005 and 2006 field trials. Additional QTL for NFNB seedling resistance to the more virulent isolate WRS858 were identified on chromosomes 2H, 4H, and 5H. A seedling resistance QTL (QRpts4) for the SFNB isolate WRS857 was detected on chromosome 4H as was a significant QTL (QRpt7) on chromosome 7H. Three QTL (QRpt6, QRpts4, QRpt7) were associated with resistance to both net blotch forms and lines with one or more of these demonstrated improved resistance. Simple sequence repeat (SSR) markers tightly linked to QRpt6 and QRpts4 were identified and validated in an unrelated barley population. The major 6H QTL, QRpt6, may provide adequate NFNB field resistance in western Canada and could be routinely selected for using molecular markers in a practical breeding program.     

Mapping of quantitative trait loci affecting Drechslera teres resistance in barley with molecular markers

 Resistance loci for seedling-stage resistance to net blotch disease (Drechslera teres) in barley were mapped with molecular markers in an F₂ population derived from a cross between the susceptible barley cultivar ‘Arena’ and the resistant Ethiopian landrace ‘Hor 9088’. Disease reactions were scored with first and second leaves of 2-week-old plants 7 and 9 days after inoculation with a single spore-derived isolate. For linkage analysis, 22 RFLP markers and 284 AFLP markers were used. The seven linkage groups covered 1153.3 cM with an average marker interval of 3.76 cM. The resistance was determined to be inherited in a quantitative manner. Altogether, 12 QTLs were mapped with positions depending on the leaf used for testing and the time period after infection. Heritability in the broad sense ranged between 0.21 and 0.37. Received: 26 May 1998 / Accepted: 9 June 1998     

Some effects of barley yellow dwarf virus on spring and winter cereals in glasshouse trials

The tolerance of spring and winter varieties of wheat, oats and barley to infection by barley yellow dwarf virus (BYDV) was examined in glasshouse tests. Severely affected plants were stunted and grain yields were considerably decreased because of decreases in both ear number and numbers and sizes of grains. Winter barley varieties were very susceptible and many were killed by BYDV infection. The winter wheat varieties were more widely tolerant than those of oats and barley. Individual seedling symptoms, although correlated with reductions in yield, could not be relied upon for accurate classification of all varieties in order of their susceptibility to infection. Symptoms of seedling infection incorporated into an index of infection permit estimates to be made o eventual decreases in yield by applying the formula DY = 1.4 × (SH+LA+LL)+18. Thus decrease in grain yield (DY) can be related to decreases in height (SH) and leaf length (LL) and increases in leaf area discoloured (LA) in seedling plants infected with BYDV.     

Die Reaktion anfälliger und resistenter Gerstensorten auf pilzliche Krankheitserreger

Response to fungus pathogen in susceptible and resistant barley cultivars IV. Water- and dry matter content
The changes in dry matter- and water content dynamics in the root and shoots of barley infected with the fungus Erysiphe graminis f. sp. hordei Marchal were studied in variously receptive cultivars. Cultivation of plants in solid (perlite) or liquid medium was compared.
The response of susceptible barley to mildew infection is characterized at First by an increase in water content and slightly in dry matter of shoots accompanied with an unsignificant decrease in root growth. However, as the fructification progressed, barley growth is reduced in all investigated parameters. The infected resistant barley indicates unsignificantly increased values in water content of shoots, only.
In the susceptible host the growth of roots in relation to shoots is markedly reduced after infection.
The percentage of water content of shoots is slightly increased in both cultivars during the first phases of infection. Then, in susceptible barley it declines, compared with the control, from the stage of full developed fructification. In the resistant barley it doesn't change more expressively. In roots of infected plants the increased hydratation is found during the whole observation period (in the resistant barley only with plants growing in liquid medium).     

The receptor-like MLO protein and the RAC/ROP family G-protein RACB modulate actin reorganization in barley attacked by the biotrophic powdery mildew fungus Blumeria graminis f.sp. hordei

Cytoskeleton remodelling is a crucial process in determining the polarity of dividing and growing plant cells, as well as during interactions with the environment. Nothing is currently known about the proteins, which regulate actin remodelling during interactions with invading pathogens. The biotrophic powdery mildew fungus Blumeria graminis f.sp. hordei (Bgh) invades susceptible barley (Hordeum vulgare L.) by penetrating epidermal cells, which remain intact during fungal development. In contrast, resistant host plants prevent infection by inhibiting penetration through apoplastic mechanisms, which require focusing defence reactions on the site of attack. We stained actin filaments in a susceptible Mlo-genotype and a near-isogenic race-non-specifically resistant barley mlo5-mutant genotype using fluorescence-labelled phalloidin after chemical fixation. This revealed that the actin cytoskeleton is differentially reorganized in susceptible and resistant hosts challenged by Bgh. Actin filaments were polarized towards the sites of attempted penetration in the resistant host, whereas when susceptible hosts were penetrated, a more subtle reorganization took place around fungal haustoria. Strong actin filament focusing towards sites of fungal attack was closely associated with successful prevention of penetration. Actin focusing was less frequent and seemingly delayed in susceptible wild-type barley expressing the susceptibility factor MLO. Additionally, single cell overexpression of a constitutively activated RAC/ROP G-protein, CA RACB, another potential host susceptibility factor and hypothetical actin cytoskeleton regulator, partly inhibited actin reorganization when under attack from Bgh, whereas knockdown of RACB promoted actin focusing. We conclude that RACB and, potentially, MLO are host proteins involved in the modulation of actin reorganization and cell polarity in the interaction of barley with Bgh.     

Resistance and susceptibility to colour forms of the aphid Sitobion avenue in spring and winter wheats (Triticum aestivum)

Glasshouse assessments of resistance to S. avenae in 29 entries of wheat and two of rye were made by releasing half-grown aphids on randomised plants at the stem extension phase of growth. Wheat cvs Kador, Amigo, Highbury and Lutescens 1377 were resistant and cvs Sentry and Talavera de Bellevue partially resistant. Cv. Klein Acero, a breeding line TB68/6/10 and Lerma Rojo selections 197 to 200 were highly susceptible to S. avenae although the latter are moderately resistant to greenbug (Schizaphis graminum) (Starks & Merkle, 1977). The rye cultivars were susceptible to 5. avenae and no cultivar was found to be resistant to Metopolophium dirhodum. Clonal stocks of S. avenae, differing in colour, varied in their ability to form large populations on susceptible cultivars, and hence in their differentiation of susceptible from resistant wheat. No clone was detected with specific ability to attack the resistant cvs Kador and Amigo.     

Effects of foliar applications of fungicides during plant growth on grain development and grain germinability in spring barley

In 1990 and 1991 grain samples of spring barley were obtained from plots in which fungal diseases were controlled until ear emergence using fungicide sprays, and from comparable plots which had received no fungicide applications. The grain was stored at 10^oC, and tested for germination at intervals during storage. Grain from plots treated with fungicide consistently exhibited more dormancy than grain from untreated plots. In 1992, grain development was monitored from anthesis to harvest-ripeness in treated and untreated plots of cvs Blenheim, Camargue and Tyne. In cvs Blenheim and Camargue, grains in fungicide-treated plots had a greater dry weight at harvest-ripeness, and dehydrated later in development than grains in untreated plots. Neither of these differences was observed in cv. Tyne. Possible interpretations of the effects of fungicide treatment of the parent plant on the physiology of the grain are discussed.     

Differential Reaction of Barley Genotypes to Rhynchosporium secalis

Spring barley cultivars currently grown in Germany are all more or less susceptible to Rhynchosporium secalis (Oudem.) J.J. Davis, but there are obvious differences in the degree of susceptibility under field conditions. Small genotypic differences may be caused by both genetic and environmental effects, respectively. To minimize the influence of environmental variation on disease expression, several inoculation methods were developed in the present study. In two experiments the effectiveness of the inoculation by spraying of single spore isolates was tested in the glasshouse and in the field, respectively. High infection levels were achieved in the glasshouse. Despite the infection of barley in the field, disease expression levels remained low due to unfavourable conditions. Another experiment showed the usefulness of infected straw applied in the autumn only for testing the seedling infection type of spring barley cultivars against R, secalis. Seedling assay scores and field infection levels were closely related (r = 0.796, P < 0.01; r = 0.911, P < 0.001). Therefore, both the spray infection technique in the glasshouse and the inoculation technique using infected straw in the field appear to be suitable to detect genetic differences in resistance/susceptibility of spring barley cultivars against leaf blotch.     

DIDYMELLA STEM-ROT OF OUTDOOR TOMATOES

In studies on Didymella stem-rot of outdoor tomatoes several possible modes of introduction of the disease were investigated, together with methods of avoiding them. The pathogen was found to invest and penetrate the seeds in diseased fruit without destroying their viability and methods were devised of ridding the seeds of infection. It was not, however, possible to demonstrate the infection of mature plants from infected seed. It was found that the disease could be introduced on contaminated canes, in potting soil and from plant debris from the previous season's crop. The spread of the disease was encouraged by overhead irrigation.
Tests of strains of various species of Lycopersicon showed that one line of L. hirsutum was highly resistant to stem and root infection although another line was highly susceptible.     

Resistance to aphids in immature wheat and barley*

Resistance to Sitobion avenue and Metopolophium dirhodum was assessed in the glasshouse by releasing immature aphids on uncovered wheat and barley plants at the tillering stage. Six cultivars of winter wheat were tested with S. avenue and of these cv. Kador was resistant and cv. Maris Huntsman susceptible. Among seven cultivars of barley, cv. CI16145 was more resistant to S. avenue than any other whilst cvs Tyra and Nirakei 10 were the most susceptible, but the cultivars did not differ significantly in the numbers of M. dirhodum on them, although there were ranking differences similar to those observed with S. avenue.     

Effects of net blotch on growth and yield of spring barley

The effect of net blotch on the growth and yield of cv. Beatrice spring barley was examined in a greenhouse experiment. Separate inoculations at growth stages 21 and 34 reduced green leaf area, root weight, leaf sheath and stem weight and tiller number. The early inoculated plants, which responded and recovered more rapidly than later treated ones, suffered a loss in grain yield and this was related to the amount of disease, the loss in green leaf area and the reduction in unit leaf rate.     

Pyrenophora teres: Population structure,virulence and aggressiveness in Southern Russia

The barley net blotch agent Pyrenophora teres (Died) Drechs. is one of the dominant fungal pathogens in agricultural crops worldwide. Here we aim to study the aggressiveness and virulence of P. teres populations collected at different ontogenesis stages (BBCH 30 and BBCH 47) from winter barley cultivars of various resistance types: moderately resistant, moderately susceptible and highly susceptible. We observed a direct proportional relationship between cultivar resistance and the aggressiveness of P. teres populations collected in both growth phases of the host plant. The isolates collected at an early stage of host plant development have a large difference in aggressiveness criteria: colony growth rate, sporulation intensity, latency period, plant damage degree, and the number of identified races. At the BBCH 30 growth stage, the growth rate of fungus colonies selected from a resistant cultivar is 1.2 times higher than that of a susceptible cultivar. The growth rate of colonies selected from resistant and susceptible cultivars in the earlier BBCH 30 stage is 1.04 times higher than the growth rate of colonies selected from the later phase. The sporulation intensity of fungal populations selected from a resistant cultivar is higher than that of populations selected from a susceptible cultivar (for BBCH 30–5.4 times, for BBCH 47–4.0 times); and it is 1.3 times higher in an earlier phase of plant development. Correlation between colony growth rate and spore formation rate in the BBCH 30 is r = 0.4. A high correlation level (r = 0.9) and notable difference between the variants were revealed when studying the duration of the latent period. The average value of plant damage by the P. teres from resistant cultivar is 4 times higher than from the susceptible cultivar in the BBCH 30 stage; and 12 times – in the BBCH 47 stage. There is a moderate negative correlation between the plant damage degree and the number of races identified from the fungal population, r = ?0.59 for the BBCH 30, r = ?0.8 for the BBCH 47. The number of races identified from P. teres populations collected in the late phase of plant growth was one third less. Our study helped to acquire new knowledge about intrapopulation processes under the influence of various factors – plant growth stage and cultivar genotype. The results obtained are the basis for the development of adaptive-integrated techniques for managing populations of the hemibiotrophic pathogen, barley net blotch.     

The effect of eyespot (Cercosporella herpotrichoides Fron.) on wheat and the influence of nitrogen on the disease

The effect of eyespot throughout the season on wheat receiving different amounts of nitrogen was studied in pot experiments. Plants inoculated in December showed chlorosis of outer leaves in February. Among plants with high nitrogen, eyespot killed 11%, caused straggling of 31% and whiteheads in 14% of the surviving ear-bearing straws, reduced yield of straw by 8% and of grain by 16%. The loss in straw yield was due to reduction in plant number, that of grain was about half due to reduction in number of ears and half to production of lighter grains. Among plants with low nitrogen the disease killed 23% of the plants, caused straggling of 86% and whiteheads in 18% of the surviving ear-bearing straws, and reduced yield of straw by 23% and of grain by 44%. The loss in straw yield was due to death of plants, that of grain was about two-thirds due to the production of fewer ears and one-third to that of lighter grains. In the high-nitrogen series the number of shoots at the time of maximum tillering was reduced by 29%; in the low-nitrogen series the disease caused reduction in height, a very uneven crop, delay in ear and anther emergence, and an increase in tail corn from 4% in the controls to 30% in the inoculated plants.
All inoculated plants became infected, but those receiving high nitrogen had only 49% of the ear-bearing straws with severe lesions at harvest, while those receiving low nitrogen had 86%. The larger number of tillers produced when nitrogen was applied may have enabled the less severely diseased shoots to survive and bear ears while the most severely infected died.     

Sensitivity of Resistance to Net Blotch in Barley

The aim of this study was to demonstrate various methods of analysing terminal net blotch, Pyrenophora teres Drechs. f. teres Smedeg., severity data from 15 spring barleys, Hordeum vulgare L., grown in Finnish official variety trials in five environments. The analyses have been developed and used principally by plant breeders for assessing crop yield, but lend themselves to use by plant pathologists. Pyrenophora teres is the major barley phytopathogen in Finland and improved resistance to it is sought. Joint regression analysis (JRA) and an additive main effects and multiplicative interaction (AMMI) model were used to investigate the data. Statistically significant genotype by environment (GE) interaction for resistance was indicated, and this included qualitative (crossover) interactions among genotypes over environments. A stable, non-sensitive, response to net blotch over environments, combined with a low mean score for terminal severity of the disease characterized the six-row barley 'Thule' which showed statistically significant crossover interaction only with 'Tyra'. 'Kustaa' exhibited the lowest mean terminal net blotch severity, but was relatively sensitive to net blotch. 'Arve' exhibited severe terminal net blotch in all environments, was relatively sensitive to environment and exhibited no crossover interaction with other genotypes. AMMI analysis appeared to represent a useful method for analysing these disease severity data, facilitating the selection of useful sources of resistance. Plots of AMMI-adjusted mean net blotch severities against first principal component axis (PCA) scores were informative for differentiating genotype response over environments, and are therefore potentially useful to plant pathologists and barley breeders seeking to gauge and subsequently improve the resistance status of barley to net blotch.     

A cDNA clone for a pathogenesis-related protein 1 from barley

A barley cDNA clone (PRb-1) corresponding to an mRNA differentially induced in resistant compared to susceptible barley cultivars by powdery mildew infection was isolated and characterised. The deduced amino acid sequence revealed 24 amino acids comprising the signal peptide and 140 amino acids of the mature peptide (15 kDa). This showed close homology to PR-1-like proteins, which have been isolated from maize, tobacco, tomato and Arabidopsis thaliana. Northern blot analysis showed accumulation of the corresponding mRNA 12 h after inoculation of resistant barley cultivars with Erysiphe graminis. Increased expression of the PRb-1 gene was also observed in resistant compared with near-isogenic susceptible barley plants following treatment with ethylene, salicylic acid, methyl jasmonate and 2,6-dichloro-isonicotinic acid.