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.     

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.     

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.     

Competence of <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> from Cruciferous Weeds and Wallflower (<Emphasis Type="Italic">Erysimum cheiri</Emphasis>) to Induce Black Rot in Cabbage

Aphanomyces euteiches Drechsler is an oomycete pathogen of leguminous crops that causes root rot, a severe disease of pea (Pisum sativum L.) worldwide. An improved understanding of the genetic structure of A. euteiches populations would increase knowledge of pathogen evolution and assist in the design of strategies to develop pea cultivars and germplasm with stable disease resistance. Twenty six primers pairs were used to amplify Sequence Related Amplified Polymorphisms (SRAP) among 49 A. euteiches isolates sampled from pea. A total of 190 polymorphic SRAP bands were generated, of which 82 were polymorphic between all the A. euteiches isolates. The percentage of polymorphic bands per primer pair ranged from 22 to 75%. According to the PIC value estimated for each marker, 60% of the SRAP markers were highly to reasonably informative (PIC > 0.25). Genetic structure of A. euteiches populations sampled in different American and French locations showed low to high genetic diversity within populations. The largest variation occurred within countries, with a total estimated genetic diversity of 0.477 and 0.172 for American and French populations, respectively. This was particularly evident from a principal component analysis (PCA) and a Minimum Spanning Networks (MSN) based on genetic profiles of isolates, which generated two different clusters, one corresponding to the French isolates and four American isolates (MV1, MV5, MV7, Ath3), and the other to American isolates. A. euteiches populations from cultivated pea in France appeared as a single unstructured population, whereas American isolates of A. euteiches diverged into three different populations.     

Pathogenicity of Aphanomyces spp. from Different Leguminous Crops in Sweden

Host range and pathogenicity of a range of Aphanomyces spp. isolates obtained from pea roots but also from a range of other field-grown leguminous crops in southern Sweden was investigated. The Aphanomyces euteiches isolates originating from pea and the few obtained isolates originating from alfalfa, green bean and yellow sweet-clover were highly pathogenic only to pea. The A. euteiches isolated from common vetch differed from these isolates by being weakly pathogenic to pea and other legumes, but highly pathogenic to common vetch. Vetch isolates also formed a well-defined separate cluster based on principal component analysis of pathogenicity pattern on tested crops. Oospores of A. euteiches were observed in root tissue of pea as well as common vetch, alfalfa, green bean, broad bean, red clover and yellow sweet-clover in the greenhouse pathogenicity tests. An Aphanomyces sp. that morphologically differed from A. euteiches, was frequently isolated from several leguminous plants, but was non-pathogenic to all tested crops in the pathogenicity tests. In isozyme analysis the banding pattern of these isolates resembled the pattern of A. cladogamus. Another, different and so far unidentified Aphanomyces sp. from roots of green bean and broad bean, was also non-pathogenic to the tested legume species. Based on the isolates tested, the results obtained suggest that the population of Aphanomyces spp. infecting legume roots in Sweden consists of a pea-specific and a vetch-specific group of A. euteiches. Two other groups comprised (i) Aphanomyces sp. isolates that resembled A. cladogamus, and (ii) isolates, which resembled neither A. euteiches nor A. cladogamus. In addition, the host range of Swedish A. euteiches isolates was not as broad as reported for A. euteiches isolates from other countries.     

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.     

Pathogenic fungi involved in root rot of peas in the Netherlands and their physiological specialization

Research on root rot pathogens of peas in the Netherlands has confirmed the prevalence ofFusarium solani, F. oxysporum, Pythium spp.,Mycosphaerella pinodes andPhoma medicaginis var.pinodella. Aphanomyces euteiches andThielaviopsis basicola were identified for the first time as pea pathogens in the Netherlands. Other pathogens such asRhizoctonia solani andCylindrocarpon destructans were also found on diseased parts of roots. F. solani existed in different degrees of pathogenicity, and was sometimes highly specific to pea, dwarf bean of field bean, depending on the cropping history of the field.A. euteiches was specific to peas, whereasT. basicola showed some degree of physiological specialization.     

Aphanomyces euteiches as a component of the complex of foot and root pathogens of peas in Dutch soils

The occurrence ofAphanomyces euteiches Drechs. in Dutch soils is reported for the first time. Isolates of the pathogen were obtained from peas (Pisum sativum L.). A bioassay was used that baited the pathogen from soil into the cortex of stem and root of seedlings of a highly susceptible pea cultivar. The pathogen could subsequently be isolated on a semi-selective medium. Screening of soil samples from 13 fields known to be infested with fungi causing foot and root rot demonstrated the presence ofA. euteiches in 10 cases. In a second screening on soil samples from 43 fields, the pathogen was present in 16 cases. A positive correlation was found between the disease severity caused byA. euteiches in the seedling bioassay and the disease severity caused by the complex of foot and root pathogens in the same soils as evidenced by a mature plant bioassay. It is considered probable thatA. euteiches has since long been a common component of the foot and root rot complex in Dutch soils but has not been detected previously due to inadequate sampling and isolation techniques.Samenvatting De aanwezigheid vanAphanomyces euteiches Drechs. in Nederlandse gronden is voor het eerst aangetoond. Isolaten van het pathogeen werden verkregen van erwten (Pisum sativum L.). De pathogene schimmel werd in petrischalen uit grond in het schorsweefsel van wortel en stengel van een zeer vatbaar erwteras gelokt. Met behulp van een semiselectief medium konden vervolgens isolaten van de schimmel worden verkregen. Toetsing van grondmonsters afkomstig van 13 percelen, waarvan bekend was dat ze besmet waren met schimmels die voetziekten in erwten veroorzaken, toonde de aanwezigheid vanA. euteiches aan in 10 gevallen. In een tweede biotoets op grondmonsters van 43 percelen bleken 16 monsters het pathogeen te herbergen. Er werd een positieve correlatie gevonden tussen de ernst van de aantasting doorA. euteiches van kiemplanten en de aantasting van volwassen planten in een biotoets in de kas. Het is waarschijnlijk dat de schimmel reeds lang in Nederlandse akkers voorkomt, maar door inadequate bemonsterings- en isolatietechnieken over het hoofd is gezien.     

Australian Uromyces viciae-fabae: Host and nonhost interaction among cultivated grain legumes

Uromyces viciae-fabae, rust of faba bean, parasitizes other legume crops such as lentils (Lens culinaris) and field peas (Pisum sativum) in some environments. In this study we examined the host range of two Australian isolates of U. viciae-fabae collected and purified from a faba bean crop and classified as U. viciae-fabae ex V. faba. Field pea (P. sativum), chickpea (Cicer arientinum), lupin (Lupinus spp.), lentil (L. culinaris), and mung bean (Vigna radiata) genotypes were tested with these isolates, as well as resistant and susceptible genotypes of the faba bean host. Race specificity for these two pathogen isolates was observed on Vicia faba, with two faba bean genotypes showing partial resistance. Both U. viciae-fabae isolates also colonized field pea seedlings and successfully produced uredinia under glasshouse conditions, despite this fungus not being known as a pathogen of Australian field pea crops. No sporulation of either isolate of U. viciae-fabae ex V. faba was observed on any of the remaining legume species tested. However, obvious differences in fungal growth were observed, ranging from small infection sites with very rare haustorium formation in mung bean to more extensive growth and the development of potential uredinial structures in chickpea. These observations are discussed in relation to the phylogenetic relationship of these host and nonhost species.     

Development of a Real-Time Polymerase Chain Reaction Assay for Quantifying Verticillium albo-atrum DNA in Resistant and Susceptible Alfalfa

ABSTRACT A precise real-time polymerase chain reaction (PCR) assay was developed for quantifying Verticillium albo-atrum DNA. The assay was used in a repeated experiment to examine the relationship between the quantity of pathogen DNA detected in infected leaves and shoots and the severity of Verticillium wilt symptoms in several alfalfa cultivars expressing a range of disease symptoms. Plants were visually inspected for symptoms and rated using a disease severity index ranging from 1 to 5, and the quantity of pathogen DNA present in leaves and stems was determined with real-time PCR. No significant differences in pathogen DNA quantity or disease severity index were observed for experiments or for cultivar-experiment interactions. Significant differences were observed between cultivars for the quantity of pathogen DNA detected with real-time PCR and also for disease severity index ratings. In both experiments, the highly resistant check cultivar Oneida VR had significantly less pathogen DNA, and significantly lower disease severity index ratings than the resistant cultivar Samauri, the moderately resistant cultivar Vernema, and the susceptible check cultivar Saranac. In both experiments, the Spearman rank correlation between the amount of V. albo-atrum DNA detected in leaves and stems with real-time PCR and disease severity index ratings based on visual examination of symptoms was positive (>0.52) and significant (P < 0.0001). These results suggest that resistance to Verticillium wilt in alfalfa is characterized by a reduced colonization of resistant genotypes by the fungus.     

Molecular quantification of the pea footrot disease pathogen (<Emphasis Type="Italic">Nectria haematococca</Emphasis>) in agricultural soils

Footrot disease due to Nectria haematococca (anamorph Fusarium solani f.sp. pisi) is an economically important disease of peas globally. However, our ability to predict accurately the likelihood of footrot infections is limited because there is no method to determine inoculum density prior to planting. In this research, a PCR-based assay was developed to quantify the pea pathogenicity gene (PEP3), exclusive to highly pathogenic forms of N. haematococca, from DNA extracted from agricultural field soils. The applicability of using quantitative PCR (qPCR) to measure this gene in soil was validated, and the relationship between PEP3 gene numbers and footrot disease was also studied. Results showed that the quantitative assay is both efficient and specific; amplification efficiency of the Q-PCR assay for the PEP3 gene was 92%. Gene copy numbers were shown to vary significantly (P = 0.01) between fields, and were positively correlated to the number of spores of pathogenic N. haematococca, and to footrot disease. PEP3 numbers of up to 100 g⁻¹ soil constituted a threshold number for infection—potentially capable of causing economically significant pea footrot disease. The density of virulent N. haematococca in soil fields capable of causing footrot disease could be determined with a high degree of accuracy, with this assay. It offers the opportunity for prediction of pea footrot infections in agricultural soils prior to cultivation.     

Host status and reaction of <Emphasis Type="Italic">Medicago truncatula</Emphasis> accessions to infection by three major pathogens of pea (<Emphasis Type="Italic">Pisum sativum</Emphasis>) and alfalfa (<Emphasis Type="Italic">Medicago sativa</Emphasis>)

Ditylenchus dipsaci, the stem nematode of alfalfa (Medicago sativa), Mycosphaerella pinodes, cause of Ascochyta blight in pea (Pisum sativum) and Aphanomyces euteiches, cause of pea root rot, result in major yield losses in French alfalfa and pea crops. These diseases are difficult to control and the partial resistances currently available are not effective enough. Medicago truncatula, the barrel medic, is the legume model for genetic studies, which should lead to the identification and characterization of new resistance genes for pathogens. We evaluated a collection of 34 accessions of M. truncatula and nine accessions from three other species (two from M. italica, six from M. littoralis and one from M. polymorpha) for resistance to these three major diseases. We developed screening tests, including standard host references, for each pathogen. Most of the accessions tested were resistant to D. dipsaci, with only three accessions classified as susceptible. A very high level of resistance to M. pinodes was observed among the accessions, none of which was susceptible to this pathogen. Conversely, a high level of variation, from resistant to susceptible accessions, was identified in response to infection by A. euteiches.     

Seed treatment of peas with fosetyl-Al against Aphanomyces euteiches

In 1983, seed dressing of peas with fosetyl-Al (Aliette) compared favourably with all other treatments on a field heavily contaminated with foot and root rot pathogens. Experiments carried out in 1983 and 1984 could not establish the reason for this phenomenon. Artificial inoculation with the most frequently isolated pathogens (Fusarium oxysporum, Phoma medicaginis var.pinodella, Pythium ultimum andFusarium solani f.sp.pisi) of sterilized soil before planting treated seeds did not reproduce the field observations, but glasshouse experiments using soil from the experimental field did. Glasshouse experiments in 1989, afterAphanomyces euteiches had been identified as a frequently occurring pea pathogen in the Netherlands, confirmed the favourable effect of fosetyl-Al as a seed treatment whenA. euteiches was in the pathogen flora.Samenvatting In 1983 bleek op een zwaar met voetrotpathogenen besmet veld zaaizaadbehandeling van erwten met fosetyl-Al (Aliette) gunstig af te steken bij alle andere behandelingen. De oorzaak kon toen niet worden gevonden. Kunstmatige besmetting van gesterilisserde grond vóór het planten van behandelde zaden met de meest geïsoleerde pathogenen (Fusarium oxysporum, Phoma medicaginis var.pinodella, Pythium ultimum enFusarium solani f.sp.pisi) gaf resultaten die niet overeenstemden met die van het proefveld, maar kasproeven met grond van het proefveld deden dat wel. Nadat was aangetoond dat het erwtepathogeenAphanomyces euteiches in Nederland veel voorkomt, bevestigden nieuwe kasproeven in 1989 het gunstige effect van fosetyl-Al als zaaizaadbehandeling wanneerA. euteiches deel uitmaakt van de pathogene bodemflora.     

Quantification of the rice false smut pathogen <Emphasis Type="Italic">Ustilaginoidea virens</Emphasis> from soil in Japan using real-time PCR

Ustilaginoidea virens is the causal agent of false smut disease of rice. In this study, we developed a real-time polymerase chain reaction (PCR) assay to clarify the relationship between false smut occurrence on rice and quantification of U. virens from soil in Japan. The method here described is sensitive, detecting less than 50 fg of pathogen DNA, and specific to the nuclear ribosomal DNA for U. virens when tested across 27 rice-pathogenic fungi and bacteria, 26 other fungi and bacteria and four plant species. As few as eight chlamydospores of U. virens per gram soil were detected when added to sterilized Gley and Ando soils. The real-time PCR assay for the soil samples was at least 100-fold more sensitive than the conventional and nested-PCR assays tested. By quantification of U. virens with real-time PCR using DNA extracted from naturally contaminated Gley soils and visual assessment of the disease in agricultural fields, a linear correlation between cycle threshold (C_T) values and the number of false smut balls was revealed. Therefore, this specific quantitative assay could be a useful tool for optimization of disease control strategies, and for studying the ecology of U. virens.     

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.     

Conventional PCR and real time quantitative PCR detection of <Emphasis Type="Italic">Phytophthora cryptogea</Emphasis> on <Emphasis Type="Italic">Gerbera jamesonii</Emphasis>

A conventional PCR and a SYBR Green real-time PCR assays for the detection and quantification of Phytophthora cryptogea, an economically important pathogen, have been developed and tested. A conventional primer set (Cryp1 and Cryp2) was designed from the Ypt1 gene of P. cryptogea. A 369 bp product was amplified on DNA from 17 isolates of P. cryptogea. No product was amplified on DNA from 34 other Phytophthora spp., water moulds, true fungi and bacteria. In addition, Cryp1/Cryp2 primers were successfully adapted to real-time PCR. The conventional PCR and real-time PCR assays were compared. The PCR was able to detect the pathogen on naturally infected gerbera plants and on symptomatic artificially infected plants collected 21 days after pathogen inoculation. The detection limit was 5 × 10³ P. cryptogea zoospores and 16 fg of DNA. Real-time PCR showed a detection limit 100 times lower (50 zoospores, 160 ag of DNA) and the possibility of detecting the pathogen in symptomless artificially infected plants and in the re-circulating nutrient solution of closed soilless cultivation systems.     

Development of PCR-based Detection Methods for the Quarantine Phytopathogen <Emphasis Type="Italic">Synchytrium endobioticum</Emphasis>, Causal Agent of Potato Wart Disease

PCR-based methods were developed for the detection and quantification of the potato pathogen Synchytrium endobioticum in soil extracts and in planta. PCR primers, based on the internal transcribed spacer region of the multi-copy gene rDNA were tested for specificity, sensitivity and reproducibility in conventional and real-time PCR assays. Soil extraction procedures compared included the Hendrickx centrifugation (HC) procedure, nested wet sieving (NWS) and a method used by the Plant Protection Service (PPS). The primers amplified a 472 bp product from S. endobioticum DNA, but did not amplify DNA from other potato pathogens, other plant pathogens, and related species. Standard cell disruption and DNA extraction and purification methods were optimized for amplification of S. endobioticum DNA from resting sporangia. DNA was successfully amplified from a single sporangium and equivalent DNA preparations from soil extracts. Low levels of target DNA in water did not amplify, possibly due to DNA loss during final purification steps. A real-time PCR assay, developed for soil-based extracts using primers and probe based on the rDNA gene sequences, involved co-amplification of target DNA along with an internal DNA fragment. Both conventional and real-time PCR methods performed well with HC- and NWS-extracts having a threshold sensitivity of 10 sporangia per PCR assay. Of the three soil extraction methods, only with the HC method could 100 g soil samples be efficiently processed in one single PCR assay. Such a high capacity assay could be useful for routine soil analysis in respect to disease risk assessments and to secure de-scheduling according to EPPO guidelines.     

Relation between cropping frequency of peas and other legumes and foot and root rot in peas

The relation between the frequency of legume crops in a rotation and the root rot severity in pea was examined in a field survey. Additionally, greenhouse experiments were performed with soil samples from legume rotation trials or from farmers' fields. The frequency of pea crops in current rotations proved to be much less than the recommended value of one in six years. The correlation between pea root rot and the number of years that pea or other legumes were not grown on the field under consideration (called crop interval) was weak. Root rot severity correlated better with the frequency of peas or legumes in general over a period of 18 years, but the frequency still explained only a minor fraction of the variation in disease index. Some experimental data pointed to the occurrence of a highly specific pathogen microflora with continuous cropping of only one legume species, but this phenomenon probably does not occur in farmers' fields. In field samples, root disease index for pea correlated well with that for field bean. The survival of resting structures of pathogens such asAphanomyces euteiches probably explains why the frequency of legume cropping has a higher impact than crop interval on root disease incidence. Pea-free periods and legume frequencies have a poor predictive value for crop management purposes.     

Development of a real-time PCR assay for detection of <Emphasis Type="Italic">Phytophthora kernoviae</Emphasis> and comparison of this method with a conventional culturing technique

Phytophthora kernoviae is a recently described pathogen causing leaf blight, aerial dieback and bleeding cankers on trees and shrubs in parts of Great Britain and Ireland and recently reported in New Zealand. This paper describes the development of a TaqMan real-time PCR assay based on internal transcribed spacer (ITS) sequence to aid diagnosis of this pathogen in culture and in plant material. The assay showed no cross reaction with 29 other Phytophthora species, including the closely related species P. boehmeriae, and detected at least 1.2 pg of P. kernoviae DNA per reaction. A rapid and simple method can be used to extract DNA prior to testing by real-time PCR, and a plant internal control assay can be used to aid interpretation of negative results. A comparison of real-time PCR and plating for 526 plant samples collected in the UK indicated that this assay is suitable for use in routine screening for P. kernoviae.     

Arbuscular Mycorrhizal Fungi Reduce Development of Pea Root-rot caused by Aphanomyces euteiches using Oospores as Pathogen Inoculum

The effects of the arbuscular mycorrhizal (AM)-fungi Glomus intraradices and Glomus claroideum on pea root-rot development caused by the pathogen Aphanomyces euteiches were investigated in a greenhouse pot-experiment, over the course of three harvests, using oospores as pathogen inoculum. Signature whole cell fatty acids 16:15c and 14:19 were used to quantify AM-fungi and A. euteiches, respectively in both roots and soil. Disease incidence was reduced in AM plants, though this effect was more pronounced in plants with G. intraradices than plants with G. claroideum, and corresponded with a greater mycorrhiza development, both intra- and extra radical in plants with G. intraradices than with G. claroideum. At the final harvest, percentage of root length with oospores was similar in roots of mycorrhizal and non-mycorrhizal plants. Despite the fact that pea root-rot development was only slightly lower in mycorrhizal plants compared to that of non-mycorrhizal plants, in terms of shoot growth and disease severity, mycorrhizal plants suffered less. This suggests a possible mycorrhiza-induced tolerance against pea root-rot. Furthermore, the degree of tolerance induction differed between the two AM-fungi included in the present study.