Susceptibility of European pear cultivars to Pseudomonas syringae pv. syringae using immature fruit and detached leaf assays

The susceptibility of thirty-three pear cultivars and two pear rootstocks to four virulent strains of Pseudomonas syringae pv. syringae was evaluated by inoculating detached immature fruits and young leaves. The four strains were similarly virulent and did not show cultivar specificity although they were isolated from different pear cultivars and exhibited different biochemical profiles. The most frequently planted pear cultivars, Conference, Abate Fetel, General Leclerc, Williams, D. Comice, El Dorado, Alexandrine, B. Anjou, Passe Crassane and the rootstock OHxF 333 were susceptible to P. syringae pv. syringae. Maximal severity values were obtained on 'Preguystar' leaves (about 90%). The rootstock Winter Nelis was less susceptible. Results with immature fruit and detached leaf assays agreed with field observations on cultivar susceptibility to bacterial blast. However, the detached leaf test gave a more accurate prediction and has the advantages that symptoms develop quickly (48 h), and leaves are available for a longer period of time than fruits. This method is proposed as a rapid and reproducible screening system of cultivar susceptibility to bacterial blast of pear.     

Specificity of polyclonal antibodies to different antigenic preparations of Pseudomonas syringae pv. pisi strain UQM551 and Pseudomonas syringae pv. syringae strain L

Polyclonal antibodies were produced against sonicated and heat-killed cells of Pseudomonas syringae pv. pisi strain UQM551 and Pseudomonas syringae pv. syringae strain L, and their specificities were compared. Evidence is presented that the serological specificity between these two pathovars lies in surface antigens. Of the surface antigens purified and tested, only flagella and lipopolysaccharide from the cell wall showed no cross-reactivity with heterologous antisera. Antisera to glutaraldehyde-fixed flagella of the two strains showed a high level of specificity. At a species or genus level, antisera prepared from heat-killed cells of P. syringae distinguished this species from all other bacterial species and genera tested, including strains of Pseudomonas fluorescens, Escherichia coli, Agrobacterium and Rhizobium.     

Interaction Between Nitrogen-Fertilized Peach Trees and Expression of syrB, a Gene Involved in Syringomycin Production in Pseudomonas syringae pv. syringae

ABSTRACT The in vitro expression of the syrB gene that controls the synthesis of syringomycin, a non-host-specific phytotoxin produced by Pseudomonas syringae pv. syringae van Hall, was studied using aqueous extracts derived from bark tissues collected from nitrogen-fertilized and nonfertilized peach trees. Expression of the syrB gene was quantified as beta- galactosidase activity expressed by P. syringae pv. syringae B3AR-132 containing a syrB::lacZ fusion. Gene expression was significantly less in three of four paired comparisons using extracts derived from fertilized versus nonfertilized trees; however, canker lengths were significantly different in only one of four comparisons. Expression was negatively correlated with plant tissue nitrogen content and positively correlated with a plant carbon/nitrogen ratio. Bark tissue from ring nematodeinfested trees had significantly higher concentrations of total soluble phenolic compounds and carbon/nitrogen ratios than bark samples from trees without nematodes, and canker size was significantly greater in trees growing in ring nematode-infested soil compared with noninfested soil. Nitrogen fertilization significantly decreased the plant carbon/nitrogen ratio, which was positively correlated with the concentration of total soluble phenolic compounds. Canker size developing after bacterial inoculation was positively correlated with higher plant carbon/nitrogen ratios and total soluble phenolic compounds. These results support the hypothesis that one reason why nitrogen fertilization decreases host susceptibility to bacterial canker is by either reducing the amount of plant metabolites that can induce syrB gene expression, or producing or increasing the concentration of compounds that antagonize syrB inducing compounds.     

Comparison of ethylene-producing Pseudomonas syringae strains isolated from kudzu (Pueraria lobata) with Pseudomonas syringae pv. phaseolicola and Pseudomonas syringae pv. glycinea

The relationships among strains of Pseudomonas syringae pv. glycinea (Psg) and Pseudomonas syringae pv. phaseolicola (Psp) isolated from kudzu ( Pueraria lobata) and bean ( Phaseolus vulgaris) were investigated. All strains tested showed a close phenotypic similarity, with the exception of the utilization of inositol and mannitol as well as the production of toxins. On this basis the strains could be divided into three groups. Group 1 consists of all strains of pathovar glycinea, group 2 includes all Psp strains isolated from kudzu, and all Psp strains isolated from bean belong to group 3. This grouping was also reflected in the genetic fingerprints using the polymerase chain reaction (PCR) with primers that anneal to dispersed repetitive bacterial sequences (rep-PCR). The rep-PCR generated fingerprints were unique for each of the three groups. The strains of group 2, Psp strains isolated from kudzu, possess certain characteristics of group 1 (ethylene production) and group 2 (phaseolotoxin production). The Psp strains from kudzu can be clearly differentiated from Psp strains isolated from bean. They utilize mannitol, produce ethylene, and are strongly pathogenic to kudzu, bean, and soybean. The results obtained show that the Psp strains from kudzu should be separated from the pathovar phaseolicola and should represent their own pathovar.     

Highly specific assays to detect isolates of Pseudomonas syringae pv. actinidiae biovar 3 and Pseudomonas syringae pv. actinidifoliorum directly from plant material

Pseudomonas syringae pv. actinidiae (Psa) is responsible for bacterial canker of kiwifruit. Biovar 3 of Psa (Psa3) has been causing widespread damage to yellow‐ and green‐fleshed kiwifruit (Actinidia spp.) cultivars in all the major kiwifruit‐producing countries in the world. In some areas, including New Zealand, P. syringae pv. actinidifoliorum (Pfm), another bacterial pathogen of kiwifruit, was initially classified as a low virulence biovar of Psa. Ability to rapidly distinguish between these pathovars is vital to the management of bacterial canker. Whole genome sequencing (WGS) data were used to develop PCR assays to specifically detect Psa3 and Pfm from field‐collected material without the need to culture bacteria. Genomic data from 36 strains of Psa, Pfm or related isolates enabled identification of areas of genomic variation suitable for primer design. The developed assays were tested on 147 non‐target bacterial species including strains likely to be found in kiwifruit orchards. A number of assays did not proceed because although they were able to discriminate between the different Psa biovars and Pfm, they also produced amplicons from other unrelated bacteria. This could have resulted in false positives from environmental samples, and demonstrates the care that is required when applying assays devised for pure cultures to field‐collected samples. The strategy described here for developing assays for distinguishing strains of closely related pathogens could be applied to other diseases with characteristics similar to Psa.     

A study of populations of Pseudomonas syringae pv. syringae on stonefruits in Victoria

In a survey of the major stonefruit nurseries in Victoria during winter 1978 and 1979, Pseudomonas syringae pv. syringae , the causal organism of bacterial canker, was found to be present on most of the stonefruit material in all nurseries but was detected most frequently on apricot.
The epiphytic populations of P.s. pv. syringae on leaves, buds and shoots of apricot and cherry were assessed periodically between 1979 and 1983 by determining the proportion of trees bearing the bacterium or by counting numbers of bacteria. Populations consistently reached peak levels during spring and late autumn, with highest levels in spring. Populations were lowest during mid- to late summer. High proportions of tree contamination and high populations coincided with periods when maximum temperatures ranged from 19° to 25°C, and when rainfall was moderately high. The significance of these findings in the light of information from other studies on the seasonal variability of host susceptibility, and in relation to chemical control, is discussed.
There was no evidence of occurrence of P.s. pv. morsprunorum in Victoria.     

Occurrence of Pseudomonas syringae pv. actinidiae on kiwifruit in Italy

Pseudomonas syringae pv. actinidiae has been isolated from kiwifruit plants for the first time in Italy. Biochemical tests were consistent with those characterizing the type-strain; pathogenicity tests yielded severe blights in the inoculated kiwifruit plants and no symptoms on lilac, pear and peach. Nutritional tests as well as whole-cell protein profiles revealed slight differences between the strains isolated in Japan and those of the present study. The main symptoms observed in the field are a red-rusty exudation covering the bark of twigs and trunks, blight of young canes and plants, angular leaf spots surrounded by chlorotic haloes and tiny cankers along the twigs.     

Characterization of Pseudomonas syringae pv. actinidiae (Psa) isolated from France and assignment of Psa biovar 4 to a de novo pathovar: Pseudomonas syringae pv. actinidifoliorum pv. nov.

Since 2008, bacterial canker of kiwifruit (Actinidia deliciosa and A. chinensis) caused by Pseudomonas syringae pv. actinidiae (Psa) has resulted in severe economic losses worldwide. Four biovars of Psa can be distinguished based on their biochemical, pathogenicity and molecular characteristics. Using a range of biochemical, molecular and pathogenicity assays, strains collected in France since the beginning of the outbreak in 2010 were found to be genotypically and phenotypically diverse, and to belong to biovar 3 or biovar 4. This is the first time that strains of biovar 4 have been isolated outside New Zealand or Australia. A multilocus sequence analysis based on four housekeeping genes (gapA, gltA, gyrB and rpoD) was performed on 72 strains representative of the French outbreak. All the strains fell into two phylogenetic groups: one clonal corresponding to biovar 3, and the other corresponding to biovar 4. This second phylogenetic group was polymorphic and could be divided into four lineages. A clonal genealogy performed with a coalescent approach did not reveal any common ancestor for the 72 Psa strains. Strains of biovar 4 are substantially different from those of the other biovars: they are less aggressive and cause only leaf spots whereas Psa biovars 1, 2 and 3 also cause canker and shoot die‐back. Because of these pathogenic differences, which were supported by phenotypic, genetic and phylogenetic differences, it is proposed that Psa biovar 4 be renamed Pseudomonas syringae pv. actinidifoliorum pv. nov. Strain CFBP 8039 is designated as the pathotype strain.     

Genetic,biochemical and pathogenic diversity of Pseudomonas syringae pv. pisi strains

Pseudomonas syringae pv. pisi is a seedborne pathogen distributed worldwide that causes pea bacterial blight. Previous characterization of this pathogen has been carried out with relatively small and/or geographically limited samples. Here, a collection of 91 strains are examined that include strains from recent outbreaks in Spain (53 strains) and from 14 other countries, and that represent all races and the new race 8, including the type race strains. This collection was characterized on the basis of 55 nutritional tests, genetic analysis (rep‐PCR, amplification of AN3 and AN7 specific markers, and multilocus sequence typing (MLST)) and pathogenicity on the differential pea cultivars to identify races. Principal component analysis and distance dendrograms confirm the existence of two genetic lineages within this pathovar, which are clearly discriminated by the AN3/AN7 markers, rep‐PCR and MLST. Strains from races 1 and 7 amplified the AN3 marker; those from races 2, 6 and 8 amplified AN7, while strains of races 3, 4 and 5 amplified either AN3 or AN7. Nevertheless, strains were not grouped by race type by any of the genetic or biochemical tests. Likewise, there was no significant association between metabolic and/or genetic profiling and the geographical origin of the strains. The Spanish collection diversity reflects the variability found in the worldwide collection, suggesting multiple introductions of the bacteria into Spain by contaminated seed lots.     

Occurrence of specific reactions induced by Pseudomonas syringae pv. syringae on bean pods, lilac and pear plants

A study on the pathogenicity of 81 strains of Pseudomonas syringae pv. syringae (PSS) isolated from 16 different hosts was conducted on lilac plants, bean pods and pear seedlings, using artificial inoculation.
Only 55 among the 81 strains induced a necrotic lesion when inoculated on lilac leaves. On bean pods, all but one of the bean isolates, and only eight strains among the 52 strains isolated from other hosts, induced typical green water-soaked lesions. On pear leaves, only pear isolates incited a typical progressive necrotic reaction, the isolates from other origins inducing no symptoms or a weak reaction limited to the inoculation point. This study indicates that in addition to the large variability observed in aggressiveness of PSS strains, host specificity occurred on bean and pear.     

Production of syringomycins and syringopeptins by Pseudomonas syringae pv. atrofaciens

All virulent strains of Pseudomonas syringae pv. atrofaciens produce in vitro substances with syringomycin-like features. All strains inhibited the growth of Geotrichum candidum in the plate assay although the extent of their growth inhibition was variable.
Purification of bioactive culture extracts of a highly virulent strain by ion exchange chromatography (Whatman CM52) yielded a main fraction which inhibited the growth of Rhodotorula pilimanae and Bacillus megaterium , and was phytotoxic to tobacco and wheat plants. In particular, the injection of this fraction in the culm of wheat plants caused phytotoxic symptoms on leaves similar to those caused in nature by the pathogen. The further purification by HPLC of the above fraction gave rise to four main bioactive substances which have been identified by spectroscopic methods (FAB-MS) and amino acid analysis as syringomycin E, syringomycin G, syringopeptin 25A and syringopeptin 25B, toxic lipodepsipeptides thus far recognized to be produced by most strains of P. syringae pv. syringae . The injection of both syringomycin E and syringopeptin 25A in wheat leaves caused necrotic symptoms; however, syringopeptin 25A was at least six times more active than syringomycin E. The possible role of the toxins in the disease development on cereals and the need for a careful examination of pathogenetic and biochemical features of P. syringae pv. atrofaciens to establish the relationships of the two pathovars in the 'syringae group' are discussed.     

上海地区大豆细菌性疫病发生危害与防治

大豆细菌性疫病又称大豆细菌性斑点病[Pseu-domonas syringaepv.glycinea(Coerper)Young,Dye&Wilkie],是我国和世界大豆产区主要病害,导致大豆产量损失和品质下降。据报道,在适宜发病条件下此病可使大豆减产18%~22%,一般减产20%左右,最高达50%以上。近年来,随着上海地区菜用大豆种植面积的不断扩大,发生了大豆细菌性疫病。为防治该病进行了田间流行调查和发生特点观测。1病害发生情况2002~2004年,对上海郊区的大豆种植地进行了调查,根据大豆细菌性疫病危害程度进行病级标准划分。0级:无症;1级:复叶上有1~5个病斑;2级:复叶上有6~10个病斑;3…     

Pathogenesis and control of bacterial speck,Pseudomonas syringae pv. tomato,on tomato*

Bacterial speck of tomato, caused by Pseudomonas syringae pv. tomato, was observed to cause severe symptoms, especially on protected tomato crops, during the winter season in the coastal area of Lebanon. This study was conducted to investigate the aetiology and pathogenesis of the bacterium involved and the efficacy of different chemicals for the control of the disease. Biochemical, physiological and pathological tests verified the identity of the bacterium involved as P. s. tomato. Periodic histological sectioning of inoculated tomato leaves showed that bacterial cells resided and multiplied in depressions and around trichome bases for 24 h before penetration through stomata and trichome basal cells. The bacteria invaded intercellular spaces and caused cell lignification, collapse and shrinkage, 48 h after inoculation. Necrotic lesions filled with bacterial masses and collapsed lignified cells were readily observable at and after 72 h. No detectable histological changes were observed in the yellow halo region surrounding the necrotic leaf specks. A thermostable toxin was produced by the pathogen and is involved in chlorotic symptom expression. An antibiotic mixture of streptomycin + oxytetracy‐cline was most effective in controlling infections followed by copper oxychloride + mancozeb, tribasic copper sulphate + sulphur, copper oxychloride and copper oxychloride + zineb.     

Sources of resistance to Pseudomonas syringae pv. phaseolicola races in Phaseolus vulgaris

One thousand and forty-eight Phaseolus bean accessions were evaluated for resistance to six races of Pseudomonas syringae pv. phaseolicola . The accessions originated from regions of the Americas and Africa where the disease is important and included wild type accessions and some known resistance sources. Resistance, graded on a five-point scale, was of two types: qualitative, which was shown to be race-specific, and quantitative. Race specific resistance genes (R-genes) were detected in 49.4% of accessions with the following gene frequencies: R1 (10.3%), R2 (0.3%), R3 (25.0%), R4 (35.0%) and R5 (0.2%).
Evidence for quantitative variation in resistance, in the absence of specific R-genes, was shown by the distribution of infection scores, 76% of accessions showing maximum susceptibility (grades 4–5), 23% showing intermediate resistance (grades 2–4), and 1% showing high levels of quantitative resistance (grades 1–2). The last 1% of accessions showed interactions which were not race-specific and it is suggested that they may possess race non-specific resistance. It is possible that several of the accessions in this category carry the recessive gene derived from PI 150414. Other accessions were of unknown parentage and may represent new sources of quantitative, potentially race non-specific, resistance. It is suggested that the combination of race specific and race non-specific resistance could provide an effective strategy for establishing durable resistance.     

Pseudomonas syringae pv. actinidiae: chemical control,resistance mechanisms and possible alternatives

Pseudomonas syringae pv. actinidiae (Psa) is a Gram‐negative bacterium that causes the bacterial canker of both green (Actinidia deliciosa) and yellow (Actinidia chinensis) fleshed kiwifruit. Since the emergence of an economically devastating Psa outbreak in Japan in the 1980s, the disease took a contagious turn causing severe economic loss to kiwifruit industries in Italy, South Korea, Spain, New Zealand and other countries. Research shows that the pathogenic strains isolated from different infected orchards vary in their virulence characteristics and have distinct genes coding for the production of different toxins. The global Psa outbreak has activated research around the world on developing efficient strategies to contain the pandemic and minimize loss to the kiwifruit industry. Chemical and biological control options, orchard management and breeding programmes are being employed in this global effort. Synergy between different disease control strategies has been recognized as important. Phytotoxicity, resistance development and regulatory measures in certain countries restrict the use of copper compounds and antibiotics, which are otherwise the mainstay chemicals against bacterial plant diseases. Therefore, because of the limitations of existing chemicals, it is important to develop novel chemical controls against Psa. Antimicrobial peptides, which are attractive alternatives to conventional antibiotics, have found promising applications in plant disease control and could contribute to expanding the chemical control tool box against Psa. This review summarizes all chemical compounds trialled so far against Psa and provides thoughts on the development of antimicrobial peptides as potential solutions for the future.