Root infection by Phytophthora cinnamomi in seedlings of three oak species

The combination of soil infestation with Phytophthora cinnamomi and repetitive flooding was studied on 1-year-old plants of Quercus ilex (holm oak) and Q. suber (cork oak). In a second experiment, using 2-year-old plants of the same species and of red oak ( Q. rubra ), the soil infestation was followed by two drought-rewatering cycles. Oak predawn leaf water potential (PLWP) and stomatal conductance (gs) were monitored during both experiments. Root infection, root loss, wilting and mortality were assessed at the end of the experiments. Q. ilex exhibited the highest susceptibility to P. cinnamomi , and Q. rubra the lowest. Root infections caused by P. cinnamomi were more severe in the flooding than in the drought experiment. The most noticeable effect of the infection on plant water relations was a decrease in stomatal conductance. This occurred at different times after inoculation, varying with species susceptibility and experiment. Inoculation with P. cinnamomi induced a decrease of PLWP in Q. ilex plants, and in some Q. suber plants exhibiting a severe root loss. The results further showed that the relationship between PLWP and gs was modified by infection with P. cinnamomi . The combination of flooding and infection with P. cinnamomi acted synergistically on the water relations of Q. ilex . By contrast, there was no significant increase in disease severity due to the postinoculation water stress imposed on the oaks.     

Phytophthora species in oak ecosystems in Turkey and their association with declining oak trees

From 1999 to 2001, a survey on the occurrence of Phytophthora spp. in the rhizosphere soil of healthy and declining oak trees was conducted in 51 oak stands in Turkey. Seven Phytophthora spp. were recovered from six out of the nine oak species sampled: P .  cinnamomi , P .  citricola , P .  cryptogea , P .  gonapodyides , P .  quercina , Phytophthora sp. 1 and Phytophthora sp. 2. The most frequently isolated species, P .  quercina , was very common on slopes susceptible to drought. It occurred in four different climatic zones and on six Quercus spp., suggesting that it is native to oaks. The second most common species, P .  citricola , was separated into three subgroups: type C was recovered only in Anatolia, whereas A and B occurred only in the European part of Turkey. Phytophthora cinnamomi was recovered at one site only, and may not be involved in oak decline in Turkey. The other four species were recovered sporadically. On affected sites there was a significant association between deteriorating crown status and the presence of Phytophthora spp., particularly P .  quercina . The occurrence of Phytophthora species was significantly influenced by soil pH. Stem inoculation tests on oak seedlings revealed that Q .  petraea was the most susceptible species.     

Variation in Pathogenicity Among South African Isolates of Phytophthora cinnamomi

Phytophthora cinnamomi isolates from South Africa were evaluated for differences in growth rate in vitro and levels of pathogenicity towards Eucalyptus smithii in the field. Inoculations were conducted in the field in summer and winter in two subsequent years at two locations in South Africa using 59 P. cinnamomi isolates. The isolates differed significantly in growth rate in vitro, as well as in levels of pathogenicity to E. smithii in the field. Growth rate in vitro was significantly influenced by interactions with culture age, geographic origin and genetic background as determined using isozymes. Levels of pathogenicity in the field were influenced by season of inoculation and average minimum temperatures at trial sites. The host from which P. cinnamomi isolates were originally obtained did not significantly affect levels of pathogenicity in the field. Culture age had a significant negative effect on growth rate in vitro and pathogenicity in the field. Significant differences in levels of pathogenicity could be found for different multilocus isozyme genotypes. Geographic origin and mating type of P. cinnamomi isolates had no significant effect on levels of pathogenicity in the field. A positive correlation was found between growth rate in vitro and levels of pathogenicity in the field. Levels of variation for pathogenicity within A1 mating type isolates were significantly lower than for A2 mating type isolates. Results of this study provide valuable information on selection of P. cinnamomi isolates for future resistance/tolerance screening assays of Eucalyptus germplasm in South Africa.     

Rapid detection of Phytophthora cinnamomi using PCR with primers derived from the Lpv putative storage protein genes

Phytophthora cinnamomi is an ecologically and economically important pathogen. In this study, PCR assays were developed with primer pair LPV2 or LPV3 for rapid detection and identification of this organism. Both primer pairs were selected from putative storage protein genes. The specificity of these primer pairs was evaluated against 49 isolates of P. cinnamomi , 102 isolates from 30 other Phytophthora spp., 17 isolates from nine Pythium spp. and 43 isolates of other water moulds, bacteria and true fungi. PCR with both primer pairs amplified the DNA from all isolates of P. cinnamomi regardless of origin. The LPV3 primers showed adequate specificity among all other species tested. The LPV2 primers cross-reacted with some species of Pythium and true fungi, but not with any other Phytophthora species. PCR with the LPV3 primers detected the pathogen at levels of a single chlamydospore or 10 zoospores in repeated tests. The PCR assay was at least 10 times more sensitive than the plating method for detection of the pathogen from artificially infested soilless medium, and, to a lesser extent, from naturally infected plants. PCR with LPV3 primers can be a useful tool for detecting P. cinnamomi from soilless media and plant tissues at ornamental nurseries, whereas the LPV2 primers can be an effective alternative for identification of this species from pure culture. Applications of these assays for detection of P. cinnamomi in other environments were also discussed.     

Evaluation of root damage to English walnut caused by five Phytophthora species

The pathogenicity of five species of Phytophthora to English walnut was studied in a greenhouse experiment. Phytophthora cinnamomi was the most aggressive species, causing severe root rot and seedling mortality. The other species tested, P. cambivora , P. citricola , P. cactorum and P. cryptogea , did not induce visible crown symptoms on seedlings 2 months after inoculation. Some strains of P. cambivora and P. cactorum also caused taproot damage to seedlings. All except one of the tested isolates caused significant necrosis of fine roots and a significant reduction of root weight compared with noninoculated seedlings. Reduction of above-ground plant development was not statistically significant. While P. cinnamomi is well known as an aggressive primary pathogen of English walnut, the other species of Phytophthora may act as predisposing factors to walnut decline, affecting root system development and increasing host vulnerability to environmental stress.     

Population Structure of Phytophthora cinnamomi in South Africa

ABSTRACT Phytophthora cinnamomi isolates collected from 1977 to 1986 and 1991 to 1993 in two regions in South Africa were analyzed using isozymes. A total of 135 isolates was analyzed for 14 enzymes representing 20 putative loci, of which four were polymorphic. This led to the identification of nine different multilocus isozyme genotypes. Both mating types of P. cinnamomi occurred commonly in the Cape region, whereas, predominantly, the A2 mating type occurred in the Mpumalanga region of South Africa. A2 mating type isolates could be resolved into seven multilocus isozyme genotypes, compared with only two multilocus isozyme genotypes for the A1 mating type isolates. Low levels of gene (0.115) and genotypic (2.4%) diversity and a low number of alleles per locus (1.43) were observed for the South African P. cinnamomi population. The genetic distance between the Cape and Mpumalanga P. cinnamomi populations was relatively low (D(m) = 0.165), and no specific pattern in regional distribution of multilocus isozyme genotypes could be observed. The genetic distance between the "old" (isolated between 1977 and 1986) and "new" (isolated between 1991 and 1993) P. cinnamomi populations from the Cape was low (D(m) = 0.164), indicating a stable population over time. Three of the nine multilocus isozyme genotypes were specific to the "old" population, and only one multilocus isozyme genotype was specific to the "new" population. Significant differences in allele frequencies, a high genetic distance (D(m) = 0.581) between the Cape A1 and A2 mating type isolates, significant deviations from Hardy-Weinberg equilibrium, a low overall level of heterozygosity, and a high fixation index (0.71) all indicate that sexual reproduction occurs rarely, if at all, in the South African P. cinnamomi population.     

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.     

The host range of isolates of downy mildew, Peronospora parasitica, from Brassica crop species

The host ranges of 33 isolates of the downy mildew fungus Peronospora parasitica from different Brassica hosts and diverse geographic origins were assessed on a standard set of Brassica accessions Isolates from each host species had a distinct host range, and in some cases isolates from the same host species but different geographic origins differed in host range. Most isolates were capable of infecting hosts other than their species of origin. These results are discussed in relation to species specificity and the conservation of certain virulence gene combinations. The epidemiological implications for cross-infection of Brassica hosts by P. parasitica isolates in the field are also considered.     

Seasonal changes in susceptibility of Quercus suber to Botryosphaeria stevensii and Phytophthora cinnamomi

Monthly inoculations of both intact plants and excised shoots of Quercus suber with the pathogenic species Botryosphaeria stevensii and Phytophthora cinnamomi were performed to investigate seasonal changes in susceptibility of this forest tree species in relation to environmental parameters and plant water status. Infection symptoms were mainly detected on seedlings inoculated from spring to autumn (April through October) with either pathogen. Mean canker sizes also showed a seasonal pattern, the higher values being recorded in the same period as above. Lesion lengths were significantly ( P  < 0·001) related to environmental minimum temperature. Mean daily minimum temperatures within the range of 5–12°C clearly inhibited lesion development of P. cinnamomi , whereas B. stevensii showed a less pronounced decrease in canker expansion at the same temperature range. In excised shoots of Q. suber inoculated monthly with B. stevensii , a negative linear relationship was found between the studied range of plant relative water content (81–91%) and canker length. In contrast, the lesions caused by P. cinnamomi were not significantly ( P  = 0·32) related to any seasonal change in water content. Some control measures for the diseases caused by both pathogens are discussed on the basis of the seasonal changes in host susceptibility observed in this study.     

重寄生菌白粉寄生孢的致病性研究

 在离体条件下,选择了6种白粉菌和49个白粉寄生孢(Ampelomyces quisqualis)菌株进行致病性测定,结果表明,白粉寄生孢菌株之间存在致病性分化且差异极显著,致病性分化主要与寄主真菌和寄主植物有关,而与地理来源相关性不大。研究同时表明,白粉寄生孢有一定的寄生专化性。     

Virulence variation of cucurbit powdery mildews in the Czech Republic – population approach

Kosman diversity models were applied to analyses of virulence (disease reaction patterns) variation of 115 isolates of two cucurbit powdery mildew (CPM) species, Golovinomyces orontii (Go) and Podosphaera xanthii (Px), collected in the Czech Republic from 2010 through 2012. Diversity within and distances between Go and Px populations and each other in a spatio-temporal context and with regard to original host plant species were analyzed based on virulence patterns of individual isolates on a set of 21 melon (Cucumis melo L.) race differential genotypes. Significant differentiation among the Go and Px pathogen populations was revealed, and the results clearly demonstrate and confirm that the set of differential C. melo genotypes is well composed because of high differentiation capacity. Differentiation of pathogens among years was significant for both species. No significant difference between Go isolates from different host plant species was established due to high variability among Go isolates, but there was significant host-specific differentiation among Px isolates. Differentiation of pathogens among regions was not detected. These results revealed high virulence variation in isolates of Go and Px, and their spatio-temporal fluctuations. High diversity in virulence of Go isolates supports the treatment of Go as a complex of different (sub)species with distinct virulence factors. Similar relationships of selected Go isolates in a UPGMA dendrogram in a previously reported multigene phylogenetic tree support the logic and suitability of the Kosman assignment based approach to population studies of organisms with asexual or mixed modes of reproduction. The approach applied in this study provides a complex view of virulence structures of powdery mildew populations, and when combined with race determination and denomination on melon, it may serve as a base to understandvirulence variation of these CPM species from a spatio-temporal viewpoint.     

葡萄根癌土壤杆菌的不同生化型对葡萄及其它植物致病性的初步研究

 从北京、内蒙古、吉林、辽宁、山东等地采到的葡萄根癌病标本中,分离到根癌土壤杆菌[Agrobacterium fumefaciens(Smith&Town.) Conn]3种生化型菌株。它们对玫瑰香葡萄的致病性明显不同。生化Ⅲ型菌株致病性强,形成大、中型瘤。生化Ⅰ型、Ⅱ型菌株致病性弱,只形成小瘤。生化Ⅲ型的不同菌株对葡萄、向日葵、烟草、番茄等8种植物的致病性也很不同。一类只能侵染葡萄,是窄寄主型;另一类除葡萄外,还能侵染向日葵、烟草、番茄,是宽寄主型。玫瑰香、龙眼等葡萄对3种生化型菌株比较敏感;莎巴珍珠、贝达等比较抗病。生化Ⅲ型和Ⅰ型的一些菌株也能侵染梨、李、杏、樱桃、榆叶梅等几种果树。从山东白雅葡萄上分离的BS33-10菌株(胭脂碱质粒类型)接种在烟草、番茄上能形成畸嵌瘤。     

Identification and pathogenicity of Cryphonectriaceae species associated with chestnut canker in China

Cryphonectriaceae species cause serious canker diseases on chestnut, oak and eucalypt trees. Recently, canker symptoms with typical orange fruiting bodies were observed on Chinese chestnut and oak trees in Hebei, Hubei, Shaanxi and Shandong Provinces in China. In the present study, isolates of these fungi were identified based on phylogenetic and morphological evidence, and their pathogenicities were tested on detached chestnut (Castanea mollissima) branches. DNA sequence comparisons of the internal transcribed spacer (ITS) regions and two regions of β-tubulin (TUB1/TUB2) indicate that these isolates represent five species in the Cryphonectriaceae, viz. Cryphonectria japonica, Cryphonectria parasitica, Aurantiosacculus castaneae sp. nov., Cryphonectria neoparasitica sp. nov. and Endothia chinensis sp. nov. The sexual morph of Aurantiosacculus is discovered for the first time and can be distinguished from the other genera in Cryphonectriaceae by dark brown ascospores and tubiform appendages at both ends. Cryphonectria neoparasitica sp. nov. is different from the other Cryphonectria species by its aseptate ascospores. Endothia chinensis sp. nov. is the sole species in Endothia infecting the host genus Castanea. Additionally, it is much smaller than E. gyrosa and narrower than E. singularis in ascospores. The inoculation results showed that these five Cryphonectriaceae fungi isolated from chestnut or oak are all pathogenic to tested chestnut branches. Cryphonectria parasitica appears to be the most aggressive fungus, followed by C. neoparasitica sp. nov., C. japonica, E. chinensis sp. nov. and A. castaneae sp. nov.     

Infection of horse chestnut (Aesculus hippocastanum) by Pseudomonas syringae pv. aesculi and its detection by quantitative real-time PCR

Pseudomonas syringae pv. aesculi is a pathogenic bacterium causing bleeding canker disease of horse chestnut ( Aesculus hippocastanum ). This is a serious disease which has been affecting horse chestnut in several European countries over the last five years; however, very little is known about the biology of the causal agent. One of the obstacles to studying this pathogen is the lengthy procedure associated with confirming its presence on the host. In this study, P. syringae pv. aesculi was isolated from lesions on different parts of horse chestnut and its pathogenicity confirmed on horse chestnut saplings using two inoculation techniques. Real-time PCR primers were developed based on gyrase B gene sequence data for the specific detection of P. syringae pv. aesculi . Primer specificity was tested on isolates of the target pathogen as well as on a broad range of related non-target bacteria and other bacterial spp. which inhabit horse chestnut. The real-time primers reliably amplified P. syringae pv. aesculi down to 1 pg of extracted DNA, with and without the presence of host DNA, and also amplified unextracted DNA in whole cells of the bacterium down to at least 160 colony forming units. Detection and quantification of the target pathogen in phloem and xylem of both naturally infected and inoculated horse chestnut tissues was also demonstrated. This quantitative real-time PCR assay provides the facility to study several important aspects of the biology of P. syringae pv. aesculi on horse chestnut including its potential for dissemination in different substrates.     

Prediction and Mapping of the Impact of Winter Temperature on the Development of Phytophthora cinnamomi-Induced Cankers on Red and Pedunculate Oak in France

ABSTRACT Phytophthora cinnamomi is the causal agent of a perennial canker that develops on the lower bole on northern red oak and pedunculate oak. The disease has a limited range in Europe, being reported only in southwest France. This limited distribution is probably linked to the susceptibility of P. cinnamomi to frost. A model was developed in previous work to estimate the impact of temperatures of <0 degrees C on the winter survival of P. cinnamomi in trunk cortical tissues and on the subsequent development of cankers. In this article, we report the use of this model to simulate canker development in 503 locations across France during a 30-year period. The predicted canker extension decreased sharply when the median P. cinnamomi winter survival index decreased from 0.95 to 0.65, with cankers that poorly developed when the median survival index was lower than 0.5 to 0.6. The actual incidence of the disease in 192 stands located across southwest France was compared with that of the model outputs. Both presence of disease in stands and frequency of cankered trees in infected stands, but not canker size on infected trees, were strongly related to the median P. cinnamomi survival index. No disease was present in stands with median survival index lower than 0.65, and the frequency of cankered trees in infected stands remained very low in stands with a median survival index between 0.65 and 0.70. Aspect was an additional factor explaining disease incidence, while the effect of elevation was likely due to its effect on winter temperatures. Maps of winter suitability to P. cinnamomi-induced cankers on oaks in France are presented.     

Genetic characterisation of <Emphasis Type="Italic">Pectobacterium wasabiae</Emphasis> causing soft rot disease of potato in New Zealand

Pectobacterium wasabiae has a narrow host range, having previously only been associated with Japanese horseradish. However, recent characterisation of Pectobacterium causing soft rotting in New Zealand has identified putative P. wasabiae isolates pathogenic to potato. In this study, phylogenetic reconstruction of acnA and mdh DNA sequences and fluorescent amplified fragment length polymorphisms (fAFLP) were used to confirm the identity of the putative P. wasabiae isolates. Both methods clustered the potato isolates closely with the type strain for P. wasabiae, ICMP9121, and also differentiated them from other plant pathogenic enterobacteria. PCR, DNA hybridisation and hypersensitive response (HR) assays were subsequently used to investigate the presence in P. wasabiae of the type III secretion system (T3SS) as well as other virulence factors known to be involved in development of disease by enterobacteria. Although all P. wasabiae strains appeared to elicit a type III-dependent HR in tobacco, genes associated with the T3SS and the putative virulence factors HecB and DspE could not be detected. Thus, genetic characterisation of P. wasabiae confirmed that it is a naturally occurring pathogen on potato, which does not possess the same suite of virulence factors that are involved in the pathogenicity of other enterobacteria on this host.     

Laboratory production of oospores of Peronospora parasitica (crucifer downy mildew) and the recovery and characterization of sexual progeny from crosses between isolates with different host specificity

The production of viable oospores of Peronospora parasitica under laboratory conditions and the recovery of isolates (referred to as sexual progeny) from these oospore populations are described. Oospores were produced when isolates of opposite sexual compatibility type, specialized to the same or different Brassica species, were grown together in seedling cotyledons of a host line capable of supporting growth of both isolates. Recovery of sexual progeny from oospore populations produced from two out of four pairings between isolates specialized to the same host species (homologous pairings) proved relatively easy. On the basis of their characterization with respect to virulence, response to phenylamide fungicides, sexual compatibility type and isoenzyme polymorphisms, there was evidence that the sexual progeny from these homologous pairings could be of hybrid origin. For the first time in a member of the Peronosporaceae, it proved possible to recover and successfully characterize a few sexual progeny from pairings between isolates specialized to different host species (heterologous pairings). However, the majority of such isolates sporulated weakly and as a consequence proved difficult to maintain and were lost. Nevertheless, some evidence for the hybrid nature of progeny from heterologous pairings was obtained.     

棉花黄萎病菌不同致病力类型在江苏省的分布

1982—1985年,对江苏省83个棉花黄萎病菌(Verticillium dahliae Kleb.)菌株进行了致病力的测定,其中致病力强的落叶型菌株13个,占测定菌株数的15.7％,主要分布在南通县恒兴乡和常熟市徐市乡、董浜乡;致病力中等的叶枯型菌株45个、致病力弱的黄斑型菌株25个,分别占测定菌株数的54.2％和30.1％,这两类菌株广泛分布江苏省各主要产棉区。试验表明棉花抗病品种“4278”是一个比较理想的鉴别寄主,对3种致病力类型的菌株的鉴别力都比较强。无论从棉花上或者从其它植物上都可以得到对棉花致病力不同的菌株,只是各类型菌株所占的比例有所不同。     

Occurrence and distribution of <Emphasis Type="Italic">Phytophthora</Emphasis> species in European chestnut stands,and their association with Ink Disease and crown decline

The Phytophthora complex associated with Castanea sativa Mill. was investigated in five European countries in 35 regions and with respect to various domestication levels. Annual precipitation and length of drought season were the main parameters that regulated the presence of Phytophthora species in the chestnut stands. Seven species of Phytophthora were detected; three of these, P. megasperma, P. cryptogea and P. syringae had not been previously reported on sweet chestnut. P. cinnamomi. P. cambivora and P. citricola were most frequently isolated. P. cinnamomi and P. cambivora were the species significantly associated with declining trees with symptoms of Ink Disease. P. cinnamomirequired distinct ecological conditions compared to the other species. P. cinnamomi was never detected in sites characterized by minimum temperatures below 1.4 °C, maximum temperature above 28 °C, or soil pH below 5.4. The results obtained provide useful information for modeling the probability of Ink Disease, crown decline and associated Phytophthora species in chestnut groves in global climatic change scenarios.     

Comparative aggressiveness of standard and variant hybrid alder phytophthoras, Phytophthora cambivora and other Phytophthora species on bark of Alnus, Quercus and other woody hosts

Pathogenicity tests were carried out on the bark of Alnus glutinosa with 19 isolates of the standard (near-tetraploid) hybrid alder phytophthora, nine isolates representing its known heteroploid variants and 11 isolates of P. cambivora , a probable parent species of the hybrid. Over a 4-year period, 12 experiments were conducted on living alder logs incubated at 20°C. Most isolates of the standard hybrid and those of the 'Dutch variant' were highly aggressive to alder bark. Isolates of the 'Swedish', 'UK' and 'German variants', and of P. cambivora , were only weakly pathogenic. Also, isolates of P. fragariae , P. cinnamomi , P . sp. 'O-group', P. cryptogea , P. megasperma , P. gonapodyides and P. citricola were either weakly or nonpathogenic. Rates of lesion development were greatest on logs cut during July–October, slower on logs cut between November and March and zero on logs cut during April, indicating a strong seasonal effect. Other evidence indicated that lesion development was subject to critical thresholds of host resistance. The standard hybrid was nonpathogenic to the bark of four other hardwood and two conifer species, indicating that it is relatively host specific. In contrast, P. cambivora was an aggressive pathogen on live bark of Quercus and Castanea . The significance of these results is discussed.