Resistance to Pseudomonas syringae in a collection of pea germplasm under field and controlled conditions

A total of 242 Pisum accessions were screened for resistance to Pseudomonas syringae pv. pisi under controlled conditions. Resistance was found to all races, including race 6 and the recently described race 8. Fifty‐eight accessions were further tested for resistance to P. syringae pv. syringae under controlled conditions, with some highly resistant accessions identified. Finally, a set of 41 accessions were evaluated for resistance to P. syringae pv. pisi and pv. syringae under spring‐ and winter‐sowing field conditions. R2, R3 and R4 race‐specific resistance genes to P. syringae pv. pisi protected pea plants in the field. Resistance sources to race 6 identified under controlled conditions were ineffective in the field. Frost effects were also evaluated in relation to disease response. Results strongly suggest that frost tolerance is effective in lowering the disease effects caused by P. syringae pv. pisi and pv. syringae under frost‐stress conditions, even in the absence of disease resistance genes, although the highest degree of this protection is reached when frost tolerance and disease‐resistance genes are combined in the same genetic background.     

A detailed evaluation method to identify sources of quantitative resistance to Fusarium oxysporum f. sp. pisi race 2 within a Pisum spp. germplasm collection

Fusarium oxysporum f. sp. pisi (Fop) is an important pathogen of field pea (Pisum sativum) worldwide. The constant evolution of the pathogen drives the necessity to broaden the genetic basis of resistance to Fop. To achieve this, it is important to have a large germplasm collection available and an accurate and efficient method for disease assessment. Here, a detailed evaluation method coupling disease incidence, disease rating over time and its related area under the disease progression curve (AUDPC) was established and used to screen a Pisum spp. germplasm collection against one isolate of Fop race 2. A large variation in the disease response of specific pea accessions ranging from highly resistant to susceptible was observed within the collection, indicating the quantitative expression of the resistance. The repetition of the inoculation experiments on a subset of 19 accessions, including two susceptible accessions, indicated that the scoring method was robust and reproducible and confirmed the highly resistant phenotypes of 11 accessions. To initiate the characterization of resistance mechanisms within these accessions, the external and internal stem symptoms were compared between these selected pea accessions, together with the extent of fungal colonization within plants. All these tests indicated that, in all resistant accessions, the resistance mechanisms efficiently stopped pathogen progression at the crown. Incorporation of these sources of resistance to breeding programmes will contribute to improved Fop resistance in pea cultivars.     

Characterization of mechanisms of resistance against Didymella pinodes in Pisum spp.

Ascochyta blight caused by Didymella pinodes is a serious disease of pea (Pisum sativum ssp. sativum) to which little resistance has been identified so far. Only incomplete resistance is available in pea germplasm although higher levels of resistance have been reported in related Pisum species. In this study we characterized histochemically the underlying resistance mechanisms in these wild species and in the pea cv. Radley, the pea cultivar with the highest level of resistance to D. pinodes. Resistance was characterized by a reduced success of colony establishment and lesion size. Histologically this was associated with higher frequency of epidermal cell death and protein cross-linking in infected epidermal cells but not with H₂O₂ accumulation and peroxidase activity.     

Benzothiadiazole and BABA improve resistance to <Emphasis Type="Italic">Uromyces pisi</Emphasis> (Pers.) Wint. in <Emphasis Type="Italic">Pisum sativum</Emphasis> L. with an enhancement of enzymatic activities and total phenolic content

Benzothiadiazole (BTH) and DL-β-aminobutyric acid (BABA) induced systemic resistance was investigated in susceptible and resistant pea genotypes against Uromyces pisi. Resistance was characterized by reduced infection frequency mainly due to decreases in appressorium formation, stomatal penetration, growth of infection hyphae and haustorium formation. Changes in β-1,3-glucanase, chitinase, phenylalanine ammonia-lyase and peroxidase activities and in total phenolics content, demonstrate that U. pisi resistance is induced by BTH and BABA treatments at early and late stages of the fungal infection process, but that the chemicals operate via different mechanisms. In fact, our study showed that BTH treatment primed the activity of pathogenesis related-proteins such as β-1,3-glucanase, chitinase and peroxidase in both susceptible and resistant genotypes. On the other hand, BABA treatment did not increase the enzymatic activities in the studied genotypes, but significantly increased their total phenolic contents.     

Characterization of the Orobanche–Medicago truncatula association for studying early stages of the parasite–host interaction

There is an increasing interest in the legume species Medicago truncatula as a model plant for structural and functional genomic studies that can be used to identify agronomically important genes in legumes. Field screening has shown very high levels of resistance to Orobanche crenata in M. truncatula. However, in vitro studies with O. crenata, Orobanche foetida, Orobanche ramosa and Orobanche minor showed useful variation among accessions at early stages of the parasite–host interaction. Significant differences were observed in the levels of germination of O. crenata and O. foetida seeds induced by different accessions of M. truncatula. Only limited germination was observed on accession SA‐4327. All accessions induced little O. ramosa and O. minor germination. Accessions also varied in the number of O. crenata and O. foetida attachments supported, with few developing on accession SA‐27774. The variation observed for induction of germination and of subsequent attachment will be useful to isolating and characterizing genes involved in the early stages of Orobanche–host plant interaction and for the study of the biosynthetic pathways of production for germination stimulants.     

Locating quantitative trait loci associated with Orobanche crenata resistance in pea

Orobanche crenata (broomrape) is a root parasite that represents the major constraint for pea (Pisum sativum) cultivation in the Mediterranean area and the Middle East. The efficacy of available control methods is minimal and breeding for O. crenata resistance is considered the most promising strategy of control. Only moderate levels of incomplete resistance are available in pea germplasm. In order to identify and map the quantitative trait loci (QTL) controlling the trait, 115 F₂ plants derived from the cross between a susceptible and a resistant parent were analysed using isozymes, random amplified polymorphic DNA and sequence tagged site markers. F₂‐derived F₃ lines were studied for O. crenata resistance under field conditions. The linkage map was constructed with MAPMAKER V2.0. Of 217 markers, 120 could be mapped into 21 linkage groups. Linkage groups consisted of 13 to two marker loci covering 1770 cM. The mean inter‐marker distance was 17.64 cM. Simple interval mapping (SIM) and composite interval mapping (CIM) were performed using QTL Cartographer. The CIM approach using five cofactors was clearly the most efficient way to locate putative QTL. Two QTL for O. crenata resistance (Ocp1 and Ocp2), explaining only a moderate portion of the observed variation (9.6% and 11.4% respectively), were detected.     

Differential Responses to Pea Bacterial Blight in Stems, Leaves and Pods Under Glasshouse and Field Conditions

Resistance to pea bacterial blight (Pseudomonas syringae pv. pisi) in different plant parts was assessed in 19 Pisum sativum cultivars and landraces, carrying race-specific resistance genes (R-genes) and two Pisum abyssinicum accessions carrying race-nonspecific resistance. Stems, leaves and pods were inoculated with seven races of P. s. pv. pisi under glasshouse conditions. For both race-specific and nonspecific resistance, a resistant response in the stem was not always associated with resistance in leaf and pod. Race-specific genes conferred stem resistance consistently, however, there was variability in the responses of leaves and pods which depended on the matching R-gene and A-gene (avirulence gene in the pathogen) combination. R2 generally conferred resistance in all plant parts. R3 or R4 singly did not confer complete resistance in leaf and pod, however, R3 in combination with R2 or R4 enhanced leaf and pod resistance. Race-nonspecific resistance conferred stem resistance to all races, leaf and pod resistance to races 2, 5 and 7 and variable reactions in leaves and pods to races 1, 3, 4 and 6.Disease expression was also studied in the field under autumn/winter conditions. P. sativum cultivar, Kelvedon Wonder (with no R genes), and two P. abyssinicum accessions, were inoculated with the most frequent races in Europe under field conditions (2, 4 and 6). Kelvedon Wonder was very susceptible to all three races, whereas P. abyssinicum was much less affected. The combination of disease resistance with frost tolerance in P. abyssinicum enabled plants to survive through the winter. A breeding strategy combining race-nonspecific resistance derived from P. abyssinicum with race-specific R-genes should provide durable resistance under severe disease pressure.     

Variation in resistance to Orobanche crenata in species of Cicer

Crenate broomrape (Orobanche crenata) is a major constraint for legume cultivation in Mediterranean agriculture. Field trials, pot and in vitro experiments demonstrated that resistance to O. crenata is present in chickpea and wild species of Cicer. The resistance is the result of the combination of several mechanisms, including low induction of parasite seed germination and in some accessions, either a darkening at the infection site on the host root that prevents establishment, or a reduced development of established parasite tubercles.     

Hydrogen peroxide scavenging mechanisms are components of Medicago truncatula partial resistance to Aphanomyces euteiches

The biochemical processes underlying the expression of resistance in the roots of Medicago truncatula against Aphanomyces euteiches infection was investigated, with emphasis on oxidative stress. The levels of H₂O₂, superoxide dismutase, peroxidase, ascorbate peroxidase, catalase, soluble phenolics and lignin were measured in the roots of two lines, A17 partially resistant and F83005.5 susceptible to A. euteiches at three infection stages; penetration of the epidermis (1 dpi), colonization of the cortex (3 dpi) and invasion of the root stele (6 dpi). A rapid and large decrease of the H₂O₂ levels in A17 roots occurred. However, in F83005.5 roots, the decrease in H₂O₂ levels was delayed until 3 dpi. In A17 roots, the activities of ascorbate peroxidase, peroxidase and catalase were induced as early as 1 dpi, whereas a general decrease in the activity of the four antioxidant enzymes was observed in F83005.5 roots. The levels of soluble phenolics and lignin were increased in A17 roots at 3 and 6 dpi, respectively. The H₂O₂ levels were negatively correlated to ascorbate peroxidase, catalase and lignin production at 1, 3 and 6 dpi, respectively in A17 roots. Physiological concentrations of H₂O₂ found in M. truncatula infected roots had no detrimental effect on the in vitro growth of this oomycete. Our data suggest that H₂O₂ does not have a direct antimicrobial effect on M. truncatula resistance to A. euteiches, but is involved in cell wall strengthening around the root stele, preventing pathogen invasion of the vascular tissues.     

Molecular and biochemical mechanisms of defence induced in pea by Rhizobium leguminosarum against Orobanche crenata

Orobanche species (broomrapes) are parasitic weeds which dramatically decrease the yields of many economically important dicotyledonous crops, including pea (Pisum sativum), in Mediterranean areas. Previously, we identified some Rhizobium leguminosarum strains, including P.SOM, which could both promote pea development and significantly reduce infection by Orobanche crenata, notably through induction of necrosis of attached parasites. In the present study, induced resistance against broomrape in the nodulated pea was shown to be associated with significant changes in rates of oxidative lipoxygenase (Lox) and phenylpropanoid/isoflavonoid pathways and in accumulation of derived toxins, including phenolics and pisatin (pea phytoalexin). Changes were followed for 5 weeks after inoculation and attack by Orobanche. In contrast to non‐inoculated plants or Orobanche only infected plants, polyphenoloxidase (PPO) activity and hydrogen peroxide content increased in response to bacteria inoculation indicating the involvement of oxidative processes. In parallel, the nodulated roots displayed high Lox activity related to the overexpression of the lox1 gene. Similarly, the expression of phenylalanine ammonia lyase (PAL) and 6a‐hydroxymaackiain 3‐O‐methyltransferase (Hmm6a) genes were induced early during nodule development, suggesting the central role of the phenylpropanoid/isoflavonoid pathways in the elicited defence. As a consequence, the derived products, phenolics and pisatin, accumulated in response to rhizobacteria and conferred mechanical and chemical barriers to the invading parasite. These results highlight the likely role of signalling pathways in induced resistance and suggest these mechanisms should be enhanced through integrated Orobanche management practices.     

Phenology and growth of Orobanche crenata Forsk (broomrape) in four legume crops

Studies were conducted in the field in 2 years comparing the phenology and growth of Orobanche crenata (Forsk) (crenate broomrape) in lentils (Lens esculenta L.) cv. Castellana, peas (Pisum sativum L.) cv. Orix, vetch (Vicia sativa L.) cv. comun and broadbean (Vicia faba L. cv. Alameda. First attachment of O. crenata to these crops took place 9–14 weeks after mid-November planting and earlier after later plantings, Differences in the first O. crenata attachment dates and in the duration of the underground growth period of O. crenata were much greater between years for any given crop than between crops in a given year: both attributes were apparently affected more by seasonal climatic conditions than by crop species, Furthermore, there was no consistent relationship found between crop growth stages and time after first attachment of the parante. This occurred in lentils and vetch while they were vegetative, in peas at late vegetative-early flowering stages, and at widely varying growth stages in broadbean, depending on planting dates and years. The maximum number of O. crenata plants successfully attached to each individual crop plant decreased in the order: peas > broadbean > lentil > vetch, with 21, 14, 10 and 8 per plant, respectively, averaged over the two seasons. Similarly, plant parasites: host dry weight ratio were 1.0, 0.7, 0.3 and 0.2 For each of these crops, respectively.     

Identification of sources of resistance to crenate broomrape (Orobanche crenata) in Spanish lentil (Lens culinaris) germplasm

The weedy root parasite Orobanche crenata constitutes a serious threat to lentils and other legumes grown in the Mediterranean and western Asia. Control strategies have centred on agronomic practices and the use of herbicides. Resistance breeding is hampered by scarcity of proper sources of resistance and of a reliable and practical screening procedure. A stepwise procedure was used here. A Spanish germplasm collection of 234 accessions of lentils (Lens culinaris) was screened for resistance to crenate broomrape under field conditions. A wide range of responses was observed, however, complete resistance was not detected. Some accessions exhibited a substantial reduction in broomrape infection. Thirty‐five lentil accessions with reduced broomrape infection were selected for further screening in the field and to determine the components of resistance in pot and in mini‐rhizotron experiments. Low infection seemed to be based on a combination of various escape and resistance mechanisms: (i) a lower root density, (ii) a lower induction of Orobanche seed germination, (iii) the reduced establishment of broomrape radicles and (iv) a limited development of established tubercles. In addition, necrosis of tubercles was observed in some accessions. This study shows how complementary in vitro screening methods may be used to rank and identify lentil accessions with potential broomrape resistance, based on different resistance mechanisms which may in the future be combined into a single cultivar by breeding.     

Resistance to powdery mildew (Blumeria graminis f.sp. avenae) in oat seedlings and adult plants

In this work, 165 Avena sativa and Avena byzantina accessions were screened for resistance to powdery mildew caused by Blumeria graminis f.sp. avenae and the defence mechanisms of resistant plants were further characterized. Ten resistant and moderately resistant accessions were selected according to macroscopic assessment. A detailed histological study of selected genotypes showed a range of defence mechanisms, acting alone or in combination, that impeded fungal development at different stages. Since the resistance observed in the collection was scarce, a study of adult plant resistance was carried out in 45 genotypes selected from field trials. Nine oat landraces and two commercial varieties showed very high levels of adult plant resistance. A detailed study of the components of the adult plant resistance revealed a high increase of penetration and post‐haustorial resistance in the fifth compared to the first leaves. Identification of the resistance sources and characterization of underlying defence mechanisms will be useful for future breeding programmes and for further cellular and molecular studies to unravel the genetic basis of resistance, in this species in particular and in cereal–powdery mildew interactions in general.     

Inheritance of resistance to <Emphasis Type="Italic">Mycosphaerella pinodes</Emphasis> in two wild accessions of <Emphasis Type="Italic">Pisum</Emphasis>

Mycosphaerella pinodes is one of the most devastating pea pathogens. Pea cultivars with adequate levels of resistance to control the disease are not so far available. However, promising levels of resistance have been identified in wild accessions of pea. In the present investigation the inheritance of resistance to M. pinodes was studied in two crosses between the susceptible pea cv. ‘Ballet’ and the partially wild resistant accessions P665 (Pisum sativum subsp. syriacum) and P42 (P. sativum subsp. sativum var. arvense). Both additive and dominant effects were important in control of resistance and susceptibility dominated over resistance.     

Differentiation of Bean pod mottle virus isolates based upon host symptoms

Accessions from Glycine, Phaseolus, and Vigna genera were screened for their reactions to different subgroups of isolates of Bean pod mottle virus (BPMV) in order to establish a differential host system. Screening results indicated that the BPMV isolates differed in pathogenic aggressiveness but not in virulence. No major resistance genes were found in soybean (Glycine max) or G. soja since all screened accessions showed mosaic or necrotic symptoms to BPMV inoculation. However, these accessions expressed differences in severity of symptoms when challenged by various BPMV isolates. The inoculation of G. tomentella accessions did not result in mosaic symptoms, and some accessions did not support systemic infection of some of the isolates. Resistance, presented as a hypersensitive reaction, was observed in some of Phaseolus and Vigna genotypes, and resistant response or susceptibility was stable to all the isolates used in the screening. In conclusion, the selected G. soja genotypes PI 407019, PI 464889A, and PI 464928, and ‘Amsoy 71’ soybean may help to separate severe (reassortant) from mild isolates of BPMV based upon their phenotypic reactions.     

四川58个小麦品种苗期抗条锈基因推导及成株期抗性表现

为了解四川小麦品种所含抗条锈基因及其在田间的抗性表现,将29个毒性谱各异的小麦条锈菌鉴别菌株于苗期接种35个已知抗条锈基因载体品系和58份四川小麦品种（系）,通过抗病谱对比分析和各品种（系）系谱分析,推导了四川小麦品种（系）的抗条锈基因型。在田间,选用中国小麦条锈菌优势小种CY32、CY33、Su-4和对Yr10、Yr24和Yr26致病的贵农22致病类型田间突变株进行混合接种,在乳熟期对各品种（系）的抗条锈性进行了鉴定。结果显示,Yr2、Yr5、Yr7、Yr9、Yr10、Yr24、Yr27、YrSpp、YrAlba等9个基因以单基因或基因组合的形式存在于44个四川小麦品种（系）中。有6个品种（系）含有Yr9基因,10个品种（系）含有Yr10基因,11个品种（系）含有Yr24基因,其中1个同时含有Yr10和Yr24基因。23个品种（系）含有未知基因及其组合。共有19个小麦品种（系）成株期对包含上述混合菌种在内的田间流行菌系表现抗病,其中5个品种抗性受成株抗性基因控制。     

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.     

Orobanche crenata (Forsk) control in Vicia faba (L.) with glyphosate as affected by herbicide rates and parasite growth stages

The broomrape (Orobanche crenata Forsk) susceptibility to glyphosate applied on faba-bean (Vicia faba L.) as affected by the parasite growth stages at the time of application was studied under field conditions. Glyphosate treatments delayed O. crenata emergence. Single glyphosate application to faba-bean infected with O. crenata predominantly at the stage (a) (small nodule) and (b) (nodules with initial vestigial roots) resulted in a moderate to low control. When the stages (c) (shoot bud already visible) or (d) (shoot and vestigial roots well developed) were the predominant stages, an excellent control was achieved with a single glyphosate application at 60 g ha^?1. Increased development stages, with the shoot emerged from the shoot bud, decreased its susceptibility to glyphosate. La lutte contre Orobanche crenata en culture de férerole avec le glyphosate; influence des doses d'herbicide et du stade de développement du parasite     

Response of rice genotypes to rice root-knot nematode (Meloidogyne graminicola) infection under varying temperature regimes

The rice root-knot nematode (RRKN), Meloidogyne graminicola, is one of the major pests of the rice–wheat cropping system. Resistance against M. graminicola in rice could be most valuable in alleviating this problem. The host response of 75 Oryza genotypes was examined at three day/night temperature regimes, 29/26°C, 34/31°C and 38/35°C, in Pluronic gel as well as in soil. Out of the 75 selected rice genotypes, only Zhenshan 97 B exhibited high resistance to this set of temperature regimes, with the least number of galls/root system. At 34/31°C, more second-stage juveniles (J2s) were hatched and J2 population densities in roots of the susceptible rice genotypes increased significantly compared with those of plants grown at the 29/26°C. In resistant genotypes, only a few J2s penetrated roots and developed into mature females. The histopathological studies revealed that in susceptible rice genotypes at high-temperature regimes, the multinucleate giant cells were well developed. The results presented in this study revealed that an increase in temperature had a significant effect on the resistance of rice genotypes and resistance appeared more pronounced in genotype Zhenshan 97 B. This resistant genotype can be used in marker-assisted selection to develop resistant elite cultivars.     

Greenhouse and field screening of wild<Emphasis Type="Italic">Lycopersicon</Emphasis> germplasm for resistance to the whitefly<Emphasis Type="Italic">Bemisia argentifolii</Emphasis>

Thirty-two accessions of wild tomato (Lycopersicon spp.) germplasm were evaluated for resistance to the whiteflyBemisia argentifolii Bellows ⇐p; Perring in a greenhouse choice bioassay. Density data were recorded for the adaxial and abaxial leaf surfaces for (i) all life stages ofB. argentifolii and (ii) types I, IV, V and VI trichomes. Individual plant selections (33 from 22 wild tomato accessions) with high resistance were subsequently tested in the field to verify the resistance found in the greenhouse screening. Resistance was defined by the density of all life stages of the whitefly observed on the eight leaflets sampled at nodes 5 and 7. Only type IV trichomes had a consistent (but low) and significant negative correlation between trichome density and whitefly density for various life stages. The highest whitefly resistance was observed inLycopersicon pennellii accessions LA 716, LA 1340 and LA 2560. The most resistantL. hirsutum f.typicum accessions were LA 1777 and LA 1353. http://www.phytoparasitica.org posting Dec. 9, 2002.