Characterization of Ascochyta isolates and susceptibility of pea cultivars to the ascochyta disease complex in Alberta

The relative virulence of 109 Ascochyta isolates collected from pea fields in Alberta from 1996 to 1998 were evaluated on 10-day-old seedlings by the excised leaf-assay technique. Twenty-eight isolates were avirulent, while the others produced lesions of various sizes on pea leaves. DNA samples from 86 isolates were amplified by the RAPD technique using PCR with single primers. One dominant genotype of Ascochyta pisi was identified throughout Alberta, but variations in virulence were not clearly differentiated by the RAPD technique. Five Ascochyta isolates, four virulent and one avirulent, were used to assess the susceptibility of 20 field pea cultivars available in Alberta, including 13 yellow types and seven green types. Based on symptom development, the yellow-type cultivars Swing, Eiffel and Delta, and the green-type cultivar Orb, were the most susceptible. Of yellow-type cultivars, Voyageur, Carneval and Montana were most resistant to A scochyta infection.     

Identification of pathovars and races of <Emphasis Type="Italic">Pseudomonas syringae,</Emphasis> the main causal agent of bacterial disease in pea in North-Central Spain,and the search for disease resistance

A total of 298 bacterial isolates were collected from pea cultivars, landraces and breeding lines in North-Central Spain over several years. On the basis of biochemical-physiological characteristics and molecular markers, 225 of the isolates were identified as Pseudomonas syringae, either pv. pisi (110 isolates) or pv. syringae (112), indicating that pv. syringae is as frequent as pv. pisi as causal agent of bacterial diseases in pea. Most strains (222) were pathogenic on pea. Further race analyses of P. syringae pv. pisi strains identified race 4 (59.1% of the isolates of this pathovar), race 2 (20.0%), race 6 (11.8%), race 5 (3.6%) and race 3 (0.9%). Five isolates (4.6%) showed a not-previously described response pattern on tester pea genotypes, which suggests that an additional race 8 could be present in P. syringae pv. pisi. All the isolates of P. syringae pv. syringae were highly pathogenic when inoculated in the tester pea genotypes, and no significant pathogenic differences were observed. Simultaneous infections with P. syringae pv. pisi and pv. syringae in the same fields were observed, suggesting the importance of resistance to both pathovars in future commercial cultivars. The search for resistance among pea genotypes suitable for production in this part of Spain or as breeding material identified the presence of resistance genes for all P. syringae pv. pisi races except for race 6. The pea cultivars Kelvendon Wonder, Cherokee, Isard, Iceberg, Messire and Attika were found suitable sources of resistance to P. syringae pv. syringae.     

Use of real-time PCR to examine the relationship between disease severity in pea and <Emphasis Type="Italic">Aphanomyces euteiches</Emphasis> DNA content in roots

Aphanomyces euteiches causes severe root rot of peas. Resistance is limited in commercial pea cultivars. Real-time fluorescent PCR assay specific for A. euteiches was used to study the relationship between disease severity and pathogen DNA content in infected peas. Five pea genotypes ranging in levels of resistance were inoculated with five isolates of A. euteiches. Plants were visually rated for disease development and the amount of pathogen DNA in roots was determined using the PCR assay. The susceptible genotypes Genie, DSP and Bolero tended to have significantly more disease and more pathogen DNA than the resistant genotypes 90-2079 and PI 180693. PI 180693 consistently had less disease, while 90-2079 had the lowest amount of pathogen DNA. The Spearman correlation between pathogen DNA quantity and disease development was positive and significant (P < 0.05) for three isolates, but was not significant for two other isolates. This suggests that the real-time PCR assay may have limited application as a selection tool for resistance in pea to A. euteiches. Its utility as a selection tool would be dependent on the correlation between disease development and pathogen DNA content for a given pathogen isolate. The accuracy and specificity of the real-time PCR assay suggests considerable application for the assay in the study of mechanisms of disease resistance and the study of microbial population dynamics in plants.     

Changes in specific virulence in Polish populations ofPhytophthora infestans: 1985–1991

Ninety-five isolates ofPhytophthora infestans collected throughout Poland during 1985–1991 and characterized for multilocus genotypes based on mating type, allozymes and DNA fingerprint, were analyzed for specific virulence to differential potato cultivars carrying ten major resistance genes. The multilocus analysis led to three groupings. The first group contained 22 isolates of a clonal lineage (PO-1) that is postulated to have been present in Europe during most of the twentieth century, but PO-1 isolates were recovered in Poland only during 1985–1988. This group contained, on average, virulence to 5.5 specific resistance genes per isolate. The second group consisted of 30 isolates in a clonal lineage (PO-4) that had not been detected before 1988. PO-4 isolates had virulence to a mean of 6.5 resistance genes per isolate. The third group was composed of 43 isolates representing 38 multilocus genotypes also not detected before 1988. These diverse genotypes had virulence to an average of 6.7 specific resistance genes per isolate. More than half (53%) of the PO-4 isolates shared a single pathotype. The group of 43 isolates was dominated by two pathotypes: the most common one (47% of the isolates) was the same pathotype that dominated PO-4 isolates; the next most common one (21%) differed from the most common one by the absence of virulence to resistance gene R5. The recent immigrant isolates (not detected before 1988) generally had virulence to a greater number of specific resistance genes than did isolates in the previous population [detected before 1988 (PO-1)]. Recent immigrant populations were dominated by one or two pathotypes, so their pathotypic diversity values were somewhat less than that of the previous population.     

Pisatin demethylation by fungal pathogens and nonpathogens of pea: association with pisatin tolerance and virulence

Previous studies have indicated that detoxification of their hosts' phytoalexins is a tolerance mechanism for some true fungi, but not the fungus-like Oomycota, and may be involved in determining the virulence of a pathogen. In the present study, the associations between demethylation of the pea phytoalexin pisatin, tolerance to pisatin, and virulence on pea were examined for 50 fungal isolates which represent 17 species of pathogens and nonpathogens of pea. All isolates ofPythium coloratum and P. irregulare failed to metabolize and were sensitive to pisatin, consistent with previous observations that members of the Oomycota generally lack the ability to metabolize and are sensitive to their hosts' phytoalexins. Among true fungi tested, the ability to demethylate pisatin was common, regardless of whether the particular isolate was pathogenic on pea or not. However, when the rate of pisatin demethylation was compared to virulence, all but one of the moderate to highly virulent isolates rapidly demethylated pisatin. In addition, the more rapidly demethylating isolates were generally more tolerant of pisatin. These results suggest that a specialized enzyme system for quickly detoxifying pisatin might be present in most pea pathogens. In previous studies a specific cytochrome P450 enzyme for demethylating pisatin was identified in the pea pathogen Nectria haematococca mating population VI, and genes (PDA genes) encoding that enzyme have been cloned from this fungus. When DNA specific for these genes was used to probe genomic DNA from other fungi that demethylate pisatin, significant hybridization was detected with only one fungus, the pea pathogen Fusarium oxysporum f. sp. pisi. If the other pea pathogens possess a specific cytochrome P450 system for detoxification of pisatin, the genes encoding these enzymes apparently share limited nucleotide similarity with N. haematococca PDA genes.     

Specific Behaviour of French Aphanomyces Euteiches Drechs. Populations For Virulence and Aggressiveness on Pea, Related to Isolates from Europe, America and New Zealand

The pathogenic variability of Aphanomyces euteiches on pea was investigated using a collection of 88 pea-infecting isolates from France and 21 isolates from Denmark, Sweden, Norway, USA, Canada and New Zealand. Aggressiveness and virulence were assessed by scoring the root symptoms on a differential set of six pea genotypes. Eleven virulence types were characterised. The virulence type I, previously described as virulent on the whole set, was predominant and included the most aggressive isolates of all geographical origins. The other types were much less prevalent, existing as one to five isolates. Three virulence types (III, IV and V) contained no French isolates. The type III, avirulent on MN313, was composed of American isolates only, and resembled the major group recently described in the USA. A wide range of aggressiveness was found within the virulence type I, and the French isolates appeared globally more aggressive than the foreign isolates. These findings indicate that isolates from the virulence type I should be used as references in breeding programs, and that pea lines PI180693 and 552 may be the most interesting resistance sources to date, despite their only partial resistance.     

Genetic diversity of the pea root pathogen Aphanomyces euteiches in Europe

The oomycete pathogen Aphanomyces euteiches causes root rot in various legume species. In this study we focused on A. euteiches causing root rot in pea (Pisum sativum), thereby being responsible for severe yield losses in pea production. We aimed to understand the genetic diversity of A. euteiches in Europe, covering a north-to-south gradient spanning from Sweden, Norway and Finland to the UK, France and Italy. A collection of 85 European A. euteiches strains was obtained, all isolated from infected pea roots from commercial vining pea cultivation fields. The strains were genotyped using 22 simple-sequence repeat markers. Multilocus genotypes were compiled and the genetic diversity between individual strains and population structure between countries was analysed. The population comprising strains from Italy was genetically different and did not share ancestry with any other population. Also, strains originating from Finland and the eastern parts of Sweden were found to be significantly different from the other populations, while strains from the rest of Europe were more closely related. A subset of 10 A. euteiches strains from four countries was further phenotyped on two susceptible pea genotypes, as well as on one genotype with partial resistance towards A. euteiches. All strains were pathogenic on all pea genotypes, but with varying levels of disease severity. No correlation between the genetic relatedness of strains and virulence levels was found. In summary, our study identified three genetically distinct groups of A. euteiches in Europe along a north-to-south gradient, indicating local pathogen differentiation.     

Further Contributions to the Development of a Differential Set of Pea Cultivars (Pisum sativum) to Investigate the Virulence of Isolates of Aphanomyces euteiches

Common root rot (Aphanomyces euteiches Drechs.) has become a very destructive disease in the French pea crops since 1993. For an accurate investigation of the virulence variability among French A. euteiches populations and between French and foreign populations, a new set of differential pea genotypes was developed. Thirty-three American and European pea lines, displaying different levels of resistance, were screened in a growth chamber against two French isolates. Symptoms (disease severity from 0 to 5, evaluating symptom surface on roots and epicotyl) and percentage of top fresh weight (inoculated/uninoculated top fresh weight ratio) were measured. From this screening 12 relatively resistant lines, from various genetic backgrounds, were identified along with a highly susceptible control. This set of 13 genotypes was inoculated under controlled conditions with 14 isolates from France, Sweden, USA, Canada and New Zealand, to investigate genotype–isolate interactions. Root symptoms were rated (disease severity), and a susceptibility/resistance threshold was established at disease severity = 1. Significant quantitative interactions were observed, and five 'resistance patterns' were identified, leading to a set of six pea genotypes: Baccara (susceptible), Capella, MN313, 902131, 552 and PI180693. Fields trials of this set in 1999 and 2000 gave the same resistance rankings than in growth chamber conditions. This set will allow more accurate assessments of the variability in virulence/aggressiveness of A. euteiches isolates from France and foreign countries, and further investigations of the epidemiological and genetic basis of pea–A. euteiches interactions.     

Variation in the resistance of barley cultivars and in the pathogenicity of Drechslera teres f. sp. maculata and D. teres f. sp. teres isolates from France

Isolates of Drechslera teres that cause net or spot-type symptoms on barley ( Hordeum vulgare ) were collected in 1986 and 1987 from fields in different regions of France. Variations in pathogenicity were evaluated using 12 barley cultivars. The Middle-Eastern cultivars Arrivate and 79-S10-10 were resistant to all isolates except R5 and S5. The Ethiopian cultivar C1 5791, previously reported to be resistant, was susceptible to the R5 and S5 biotypes. There was a high correlation coefficient between the classification of cultivars for resistance to D. teres f. sp. teres and D. teres f. sp. maculata. A method for conserving the virulence of the isolates on straw is evaluated. The virulence level of the isolates was the same after 4 years of storage using this method.     

江西省稻瘟病菌毒性的群体结构分析

用江苏省的吴江、赣榆、通州、高邮和宜兴等5个代表地区的324个稻瘟病菌株接种13个日本已知抗病基因品种，结果表明，江苏省稻瘟病菌对携带有Pi-Ka,Pi-ta,Pi-ta2和Pi-sh抗病基因品种的可致病比例较高，毒力频率达64．2％－97．2％，对携带有Pi-iPi-z和Pi-b抗病基因的品种毒力频率较低，在0－11．9％之间；江苏省稻瘟病菌毒性的群体结构年度间有一定变化，表现为对某些抗病基因品种毒力频率的明显上升或下降。从江苏省10年主栽品种上分离的稻瘟病菌株对13个已知抗病基因品种的毒力频率测定结果表明，不同类型主栽品种上分离的菌株其毒性结构组成有一定差异,表明江苏省水稻主栽品种的抗性类型存在一定差异。     

Durability of resistance in lily to basal rot: evaluation of virulence and aggressiveness among isolates ofFusarium oxysporum f. sp.lilii

Prospects of durability of resistance in lily to basal rot have been evaluated by testing the virulence and aggressiveness of 31 isolates ofFusarium oxysporum f. sp.lilii towards a number of different resistance sources inLilium spp. Isolates differed strongly in aggressiveness as did species and cultivars ofLilium spp. in resistance. Significant interactions were observed between isolates of the pathogen and genotypes ofLilium spp., but the magnitude was very small compared to the main effects. The interactions were mainly due to a small group of isolates with low aggressiveness. It is argued that the interactions might be based on minor genes. No major break down of the resistance was found. For practical purposes it will be sufficient to use highly aggressive isolates in screening tests.     

Competitiveness against grass weeds in field pea genotypes

Summary Field pea is an important pulse crop in southern Australia, but its competitiveness against weeds is low. Two experiments were conducted in consecutive years to examine the ability of different genotypes to compete against grass weeds. Field pea was grown in the presence or absence of Lolium rigidum or wheat. In each experiment significant genotypic differences in field pea competitiveness occurred and some genotypes had consistently high or low levels of competitiveness in both years. Tall genotypes generally suppressed L. rigidum and wheat more effectively than short genotypes. Leaf type (conventional, semileafless or tare-leaf) only affected competitiveness before flowering and had no effect on yield loss from weed competition. Maturity had little effect on competitive ability. A genotype × environment analysis for competitive ability showed that there was considerable variation in the response to the level of competition (environment) as well as in mean competitiveness among field pea genotypes. Using wheat as a weed identified genotypic differences in field peas more effectively and more consistently than using L. rigidum . Visual assessment of wheat growth with field pea was strongly correlated with a specific genotype's competitiveness, which may form the basis of a simple field-based screening method for competitive ability.     

The genetic control and expression of specificity in Bremia lactucae (lettuce downy mildew)

The inheritance of specific virulence in Bremia lactucae was studied in crosses involving 12 heterothallic isolates of the fungus. In one cross, virulence to eight of the II specific resistance factors examined segregated in the F₁ generation. Although there were exceptions, most of the data were consistent with the hypothesis that pathogenicity was controlled by independent single loci with avirulence dominant to virulence. Linkage between loci determining virulence on R2 and R11 was confirmed. Loci controlling virulence on R5 and R8 also appeared to be linked but these may be identical R-factors. Contrary to a previous suggestion, the locus determining virulence to R10 was independent of that for R5/R8. The expression of virulence to three R-factors (Rl, R4 and R5/R8) was influenced by independent second loci. The presence of a dominant allele at the second locus inhibited avirulence. The expression of avirulence on R6 seemed to be influenced by modifier genes and environment in some isolates. Although the cultivars Mildura, Bourguignonne, Sucrine and Captain were originally thought to contain a single resistance factor, these data suggest that Sucrine carries R5/R8 in addition to R10 whilst Mildura may carry Rl in addition to R3. The stock of Bourguignonne appeared to be a mixture of resistance genotypes. The data add additional support to the suggestion that Capitan (Rll) may carry two resistance factors.     

Genetic relationship between races of Pseudomonas syringae pv.pisi and cultivars of Pisum sativum

Isolates of Pseudomonas syringae pv. pisi from the UK and overseas were categorized into six races on the basis of their reactions to a range of differential pea (Pisum sativum) cultivars. Race 2 was predominant among the isolates examined and this probably reflects its relative international importance. A previously uncharacterized race (race 6) was virulent on all cultivars tested. Resistance to races 1-5 was widespread in commercial cultivars and breeding lines with more than 75% showing resistance to one or more races. A preliminary study of the inheritance of resistance indicated that for races 1, 2 and 3, resistance was controlled by different dominant genes. The genetic basis for the relationship between races of P. syringae pv. pisi and pea cultivars was explained in terms of a gene-for-gene relationship involving five matching gene pairs. With further clarification of the genetics of resistance this host-pathogen association will meet most of the requirements of a model system for the study of the genetic and molecular basis of pathogenicity and host specificity.     

Reaction of genotypes from several species of grain and forage legumes to infection with a French pea isolate of the oomycete <Emphasis Type="Italic">Aphanomyces euteiches</Emphasis>

The susceptibility/resistance to Aphanomyces euteiches of various genotypes (cultivars and breeding lines) of several grain legume species was assessed in controlled conditions. A total of 279 genotypes from the major grain legumes grown in temperate climates (faba bean, chickpea, lentil, lupin and common vetch) and three other legumes frequently cultivated in France (French bean, clover and alfalfa) were screened with one pea-infecting isolate from France. Four different categories of susceptibility/resistance were identified among the legume species/cultivars tested with the pea A. euteiches isolate: (1) susceptible legume species (lentil, alfalfa, French bean) among which low levels of partial resistance was observed; (2) legume species including susceptible genotypes and genotypes with high levels of resistance (common vetch, faba bean and clover), (3) species with a very high level of resistance (chickpea) and (4) species displaying no symptoms (lupin). It is therefore important to consider pathogen-species and pathogen-genotype interactions when defining the host specificity of A. euteiches and considering the possible role of different legume species in increasing or decreasing the soil inoculum potential.     

Resistance in Australian barley (Hordeum vulgare) germplasm to the exotic pathogen Puccinia striiformis f. sp. hordei,causal agent of stripe rust

Eighty‐eight Australian and 10 international barley cultivars were assessed for resistance to the barley stripe (yellow) rust pathogen, Puccinia striiformis f. sp. hordei (Psh). All cultivars were tested for seedling resistance to two UK‐derived isolates of Psh (11.01 and 83.39) that were shown to differ in virulence based on responses on 16 differential barley genotypes. The 98 barley cultivars differed substantially in stripe rust response; 45% were susceptible to Psh 11.01, 53% to Psh 83.39 and 44% to both isolates. The observed diverse infection types (ITs) suggest the presence of both known and uncharacterized resistance. However, further multipathotype tests are required for accurate gene postulation. The Yerong × Franklin (Y×F) doubled haploid (DH) population was phenotypically assessed as seedlings using both Psh isolates. Yerong and Franklin were immune and highly resistant, respectively, to both isolates used in this study. Marker‐trait and QTL mapping identified a major effect on the long arm of chromosome 7H contributed by Franklin in response to all isolates. The resistance of Yerong was mapped to 113·96 and 169·38 cM on chromosome 5HL in response to Psh 11.01 and 83.39, respectively. The Psh resistance sources identified in this study can be used for further genetic analysis and introgression for varietal improvement.     

Host Specificity of Phytophthora infestans on Tomato and Potato in Ecuador

ABSTRACT Sixty Ecuadorian isolates of Phytophthora infestans from potato and 60 isolates from tomato were compared for dilocus allozyme genotype, mitochondrial DNA haplotype, mating type, and specific virulence on 11 potato R-gene differential plants and four tomato cultivars, two of which contained different Ph genes. Restriction fragment length polymorphism (RFLP) fingerprints of subsamples of isolates from each host were compared by using RG57 as the probe. All potato isolates had the allozyme genotype, haplotype, and mating type of the clonal lineage EC-1, which had been previously described in Ecuador. With the same markers, only one isolate from tomato was classified as EC-1; all others belonged to the globally distributed US-1 clonal lineage. RFLP fingerprints of isolate subsets corroborated this clonal lineage classification. Specific virulence on potato differentials was broadest among potato isolates, while specific virulence on tomato cultivars was broadest among tomato isolates. Some tomato isolates infected all tomato differentials but no potato differentials, indicating that specific virulence for the two hosts is probably controlled by different avirulence genes in P. infestans. In two separate experiments, the diameters of lesions caused by nine isolates from potato and 10 from tomato were compared on three tomato and three potato cultivars. All isolates produced larger lesions on the host from which they were isolated. No isolates were found that were highly aggressive on both tomato and potato. We conclude that there are two different populations of P. infestans in Ecuador and that they are separated by host.     

Isolation, identification and molecular characterization of physiological races of apple powdery mildew (Podosphaera leucotricha)

The development of apple varieties displaying durable resistance against powdery mildew is one of the major aims in apple breeding programmes worldwide. For a reliable judgment of the resistance of different Malus genotypes, an extended knowledge about the virulence of the pathogen is necessary. To prove the existence of physiological races of Podosphaera leucotricha , 31 monoconidial isolates of the obligate biotrophic fungus representing five locations within Europe have been established and maintained over a period of 3–4 years. The isolates were maintained on in vitro shoots of the highly susceptible apple cv. Gibb's Golden Gage. An AFLP-based DNA fingerprinting protocol was developed and, using 54 stably reproducible AFLP markers, a dendrogram revealed genetic variability among different isolates of P. leucotricha . Although the molecular characterization of the isolates showed an overall low level of genetic variability, the high phenotypic diversity among European isolates suggest that sexual reproduction may also be involved in the disease cycle of the pathogen in Europe. Phytopathological tests using detached leaves of a collection of 36 Malus genotypes allowed the differentiation of five selected isolates by their virulence patterns. A high level of diversity in terms of virulence was obtained in P. leucotricha. From the present study, based on apple breeding germplasm, cultivars and Malus species, it can be concluded that physiological races of P. leucotricha do indeed exist in Europe.     

The role of aggressiveness and competition in the selection of Phytophthora infestans populations

Selection within populations of Phytophthora infestans was investigated by comparing the aggressiveness of single‐lesion isolates on detached leaflets of four potato cultivars with differing levels of race‐nonspecific resistance to P. infestans. The isolates included 23 representative of Northern Ireland genotypes from the early 2000s, used to inoculate previously reported field trials on competitive selection (2003–2005), plus 12 isolates recovered from the 2003 trial. The cultivars were those planted in the previous trials: Atlantic (blight‐susceptible) and Santé, Milagro and Stirling (partially resistant). Very highly significant variation for latent period, infection frequency and lesion area was found between genotypes and cultivars; differences between genotypes were more marked on the more resistant cultivars, but no one genotype was the most aggressive across all. Detached leaflets were also inoculated with mixtures of isolates from each genotype group at three sporangial concentrations: differences in aggressiveness between genotypes were more apparent at lower concentrations and on the more resistant cultivars. Genotype groups that were the most aggressive on the more resistant cultivars tended to be those selected by the same cultivars in the field. A mixture of all isolates of all genotypes was used to inoculate detached leaflets of the same cultivars. With one exception, single spore isolates recovered from any one leaflet belonged to a single genotype, but different genotypes were recovered from different cultivars. Phytophthora infestans isolates from Northern Ireland showed significant variation for foliar aggressiveness, and pathogen genotypes exhibited differential aggressiveness to partially resistant cultivars and interacted competitively in genotype selection.