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1.
中国猕猴桃细菌性花腐病菌的鉴定   总被引:4,自引:0,他引:4  
 从福建、湖南和湖北猕猴桃病花上分离到能引起花腐病的32个细菌菌株,经细菌学和BiologGN测试板测定,可以看出中国的猕猴桃细菌性花腐病菌与新西兰的猕猴桃花腐病菌、丁香假单胞菌丁香致病变种Pseudomonas syringae pv.syringae和绿黄假单胞菌P.viridiflava相似,与萨氏假单胞菌P.savastanoi和猕猴桃溃疡病菌P.syringae pv.actinidiae有更多的不同,但是DNA/DNA同源性测定结果却显示出中国的菌株可分为2个类型:第1个类型与新西兰猕猴桃花腐病菌和萨氏假单胞菌有很高的同源性,第2类型与绿黄假单胞菌有很高的同源性,说明中国菌株分别属于这2个种。第1类型来自于福建和湖北,第2类型来自于湖南。  相似文献   

2.
ABSTRACT Pseudomonas syringae pv. actinidiae, the causal agent of canker in kiwifruit (Actinidia spp.) vines, was first detected in Japan in 1984, followed by detections in Korea and Italy in the early 1990s. Isolates causing more severe disease symptoms have recently been detected in several countries with a wide global distribution, including Italy, New Zealand, and China. In order to characterize P. syringae pv. actinidiae populations globally, a representative set of 40 isolates from New Zealand, Italy, Japan, South Korea, Australia, and Chile were selected for extensive genetic analysis. Multilocus sequence analysis (MLSA) of housekeeping, type III effector and phytotoxin genes was used to elucidate the phylogenetic relationships between P. syringae pv. actinidiae isolates worldwide. Four additional isolates, including one from China, for which shotgun sequence of the whole genome was available, were included in phylogenetic analyses. It is shown that at least four P. syringae pv. actinidiae MLSA groups are present globally, and that marker sets with differing evolutionary trajectories (conserved housekeeping and rapidly evolving effector genes) readily differentiate all four groups. The MLSA group designated here as Psa3 is the strain causing secondary symptoms such as formation of cankers, production of exudates, and cane and shoot dieback on some kiwifruit orchards in Italy and New Zealand. It is shown that isolates from Chile also belong to this MLSA group. MLSA group Psa4, detected in isolates collected in New Zealand and Australia, has not been previously described. P. syringae pv. actinidiae has an extensive global distribution yet the isolates causing widespread losses to the kiwifruit industry can all be traced to a single MLSA group, Psa3.  相似文献   

3.
The genetic diversity of 71 Pseudomonas savastanoi pv. savastanoi strains isolated from different host species and from diverse geographical regions was determined by fluorescent amplified fragment length polymorphism (f-AFLP) analysis. The study was carried out using three different selective primer combinations. Strains of P. syringae pv. syringae , P. syringae pv. phaseolicola , P. syringae pv. glycinea , P. syringae pv. tagetis and P. amygdali were also included as outgroups. Based on cluster analysis of f-AFLP data, all P. savastanoi pv. savastanoi strains showed a high degree of similarity, grouping in a cluster and forming a taxon clearly separate from outgroup strains. AFLP analyses failed to support placing strains of P. savastanoi pv. savastanoi , P. syringae pv. phaseolicola and P. syringae pv. glycinea in the same species. Strains of P. savastanoi pv. savastanoi formed subclusters that correlated with the host species. Strains identified within these subclusters were related to the geographical region where the strains were isolated. Strains of P. savastanoi pv. savastanoi from olive were divided into two subclusters. Strains from oleander were differentiated from those from ash and were divided into two additional subclusters, distinct from olive strains. Three strains isolated from jasmine showed a high level of similarity among them but, at a lower Dice similarity coefficient, were linked to a subcluster including olive strains. Finally, two strains isolated from privet were similar to strains from olive and were included in the same subcluster.  相似文献   

4.
猕猴桃溃疡病菌的分子检测技术研究   总被引:3,自引:0,他引:3  
 猕猴桃溃疡病是猕猴桃生产上的主要病害,为建立该病的快速诊断技术,本实验通过RAPD分析获得一条1 300 bp左右的致病菌的特异片段,对该片段进行克隆测序,在测序的基础上设计并合成一对特异引物F7/R7,优化特异引物扩增条件,并验证引物的特异性和灵敏性。利用该特异引物对包括猕猴桃溃疡病菌在内的14个菌株基因组DNA进行PCR扩增表明,只有猕猴桃溃疡病菌能扩增出1条约为950 bp的特异条带,其他菌株及对照均未扩增出特异条带。对采自果园的染病枝干组织和接种致病菌的枝干组织的检测表明,该特异引物能特异性地检测到猕猴桃溃疡病菌的存在,其在组织中的检测灵敏度为100 fg/μL。因此,利用设计合成的特异引物F7/R7,参考优化的体系和程序,结合简单的试剂盒法提取猕猴桃溃疡病菌或植物组织DNA,可以在短时间内完成对该病原菌的分子检测。  相似文献   

5.
The bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae is a severe threat to kiwifruit production worldwide. Many aspects of P. syringae pv. actinidiae biology and epidemiology still require in-depth investigation. The infection by and spread of P. syringae pv. actinidiae in xylem and phloem was investigated by carrying out artificial inoculation experiments with histological and dendrochronological analyses of naturally diseased plants in Italy. We found that the bacterium can infect host plants by entering natural openings and lesions. In naturally infected kiwifruit plants, P. syringae pv. actinidiae is present in the lenticels as well as in the dead phloem tissue beneath the lenticels, surrounded by a lesion in the periderm which appears to indicate the importance of lenticels to kiwifruit infection. Biofilm formation was observed outside and inside plants. In cases of advanced stages of P. syringae pv. actinidiae infection, neuroses of the phloem occur, which are followed by cambial dieback and most likely by infection of the xylem. Anatomical changes in wood such as reduced ring width, a drastic reduction in vessel size, and the presence of tyloses were observed within several infected sites. In the field, these changes occur only a year after the first leaf symptoms are observed suggesting a significant time lapse between primary and secondary symptoms. It was possible to study the temporal development of P. syringae pv. actinidiae-induced cambial dieback by applying dendrochronology methods which revealed that cambial dieback occurs only during the growing season.  相似文献   

6.
猕猴桃溃疡病菌在中国的适生性分析   总被引:2,自引:0,他引:2  
通过分析猕猴桃溃疡病菌在中国的适生性,为科学制定有效的检疫监管措施,防范其入侵和扩散,确保猕猴桃产业健康发展提供理论依据。本研究根据前人研究结果,采用模糊数学综合评判的原理和方法,定量分析猕猴桃细菌性溃疡病菌(Pseudomonas syringae pv.actinidiae)在我国各个地区的适生性。猕猴桃溃疡病菌在我国最适宜的省份主要分布在四川、云南、贵州、福建、安徽、湖南、湖北、河南、江西、陕西、浙江、重庆、西藏。鉴于该病具有发生发展迅速,危害性强,防治难度大等特点,应当加强猕猴桃种苗等繁殖材料的检疫,加强对果园的管理和病害监测,积极采取有效的防治措施并加强抗病育种方面的研究。  相似文献   

7.
Pseudomonas syringae pv. persicae , the pathogen causing bacterial decline of stone-fruit, was first noted almost simultaneously in France and in New Zealand, together with a closely related pathogen from myrobalan plum in England. The relative similarity of 31 strains from these three countries was examined by comparing DNA restriction endonuclease fragment patterns. Fragment patterns produced from digested DNA samples which were electrophoresed on polyacrylamide gels and visualized by silver-staining, were analysed using the software package Gelcompar. The fragment patterns produced by strains from France and England formed homogeneous but separate groups, while those from New Zealand were relatively heterogeneous. This finding suggests that the New Zealand population of P.s. persicae is older than those found in Europe. Problems in explaining the distribution of the pathogen are discussed.  相似文献   

8.
This study assessed the use of in vitro olive plants to evaluate the virulence of Pseudomonas savastanoi pv. savastanoi strains isolated from olive and P. savastanoi pv. nerii strains isolated from oleander knots. First, different olive isolates were inoculated into stem wounds and differences in knot formation and weight of overgrowths were observed for the selected strains. Tissue proliferation was clearly visible in all inoculated plants 30 days after inoculation. Virulence of P. savastanoi pv. nerii mutants with defects in regard to biosynthesis of indole-3-acetic acid and/or cytokinins was tested using this system. In agreement with data previously reported, all mutant strains multiplied in olive but induced attenuated symptoms. To analyze the virulence of P. savastanoi pv. savastanoi affected in their ability to grow in olive tissue, a trpE tryptophan auxotroph mutant was generated using a collection of signature tagged mutagenesis transposons. Virulence of this mutant was clearly reduced as evidenced by swelling of the olive tissue that evolved into attenuated knots. Furthermore, mixed infections with its parental strain revealed that the wild-type strain completely out-competed the trpE mutant. Results shown here demonstrate the usefulness of in vitro olive plants for the analysis of P. savastanoi pvs. savastanoi and nerii virulence. In addition, this system offers the possibility of quantifying virulence differences as weight of overgrowths. Moreover, we established the basis for the use of mixed infections in combination with signature tagged mutagenesis for high-throughput functional genomic analysis of this bacterial pathogen.  相似文献   

9.
ABSTRACT The virulence of Pseudomonas syringae subsp. savastanoi, which causes hyperplastic symptoms (knots) on olive plants, is associated with secreted phytohormones. We identified a Tn5-induced mutant of P. syringae subsp. savastanoi that did not cause disease symptoms on olive plants although it was still able to produce phytohormones. In addition, the mutant failed to elicit a hypersensitive response in a nonhost plant. Molecular characterization of the mutant revealed that a single Tn5 insertion occurred within an open reading frame encoding a protein 92% identical to the HrcC protein of P. syringae pv. syringae. Moreover, sequence analysis revealed that the gene encoding the HrcC protein in P. syringae subsp. savastanoi was part of an operon that included five genes arranged as in other phytopathogenic bacteria. These results imply that hrp/hrc genes are functional in P. syringae subsp. savastanoi and that they play a key role in the pathogenicity of this plant pathogen.  相似文献   

10.
Occurrence of Pseudomonas syringae pv. actinidiae on kiwifruit in Italy   总被引:2,自引:0,他引:2  
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.  相似文献   

11.
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.  相似文献   

12.
Several published polymerase chain reaction (PCR) primers to identify Pseudomonas syringae pv. actinidiae, the causal organism of bacterial canker of kiwifruit, were found not to be specific. Two new sets of PCR primers, PsaF1/R2 and PsaF3/R4, were designed to be complementary to a portion of the 16S–23S rDNA intertranscribed spacer (ITS) regions. These primers amplified a DNA fragment from strains of P. syringae pv. actinidiae, but not from 56 strains of bacteria from six genera and 17 species, except for a strain of the tea pathogen, P. syringae pv. theae. When tested against DNA extracted from a further 20 strains from Japan, Korea, Italy and the USA deposited in culture collections as P. syringae pv. actinidiae, all except six cultures produced the expected product of 280 bp with PsaF1/R2 and 175 bp with PsaF3/R4. Results of multilocus sequence analysis using five housekeeping genes (gyrB, acnB, rpoD, pgi and cts) showed that none of these six strains was phylogenetically similar to P. syringae pv. actinidiae. In contrast to the P. syringae pv. actinidiae type strain, these strains were positive in the determinative tests for ice nucleation and syringomycin production. It is suggested that these six strains were incorrectly identified as P. syringae pv. actinidiae. It was not possible to distinguish P. syringae pv. actinidiae from the phylogenetically similar P. syringae pv. theae using the ITS, gyrB, acnB, rpoD, pgi or cts gene regions to design PCR primers. Because P. syringae pv. theae is unlikely to be found on kiwifruit, primers PsaF1/R2 and PsaF3/R4 are recommended for screening bacteria isolated from kiwifruit tissue.  相似文献   

13.
ABSTRACT Pseudomonas savastanoi pv. savastanoi causes olive knot disease, which is present in most countries where olive trees are grown. Although the use of cultivars with low susceptibility may be one of the most appropriate methods of disease control, little information is available from inoculation assays, and cultivar susceptibility assessments have been limited to few cultivars. We have evaluated the effects of pathogen virulence, plant age, the dose/response relationship, and the induction of secondary tumors in olive inoculation assays. Most P. savastanoi pv. savastanoi strains evaluated were highly virulent to olive plants, but interactions between cultivars and strains were found. The severity of the disease in a given cultivar was strongly dependent of the pathogen dose applied at the wound sites. Secondary tumors developed in noninoculated wounds following inoculation at another position on the stem, suggesting the migration of the pathogen within olive plants. Proportion and weight of primary knots and the presence of secondary knots were evaluated in 29 olive cultivars inoculated with two pathogen strains at two inoculum doses, allowing us to rate most of the cultivars as having either high, medium, or low susceptibility to olive knot disease. None of the cultivars were immune to the disease.  相似文献   

14.
ABSTRACT Strains of Pseudomonas syringae (78 strains and 43 pathovars) and other strains (79) of plant and insect origin were examined for the presence of the ethylene-forming enzyme gene (efe) by polymerase chain reaction (PCR) assay. The sequence of the efe gene of P. syringae pv. phaseolicola PK2 was used to design two primer sets for amplification of the gene. In addition to P.syringae pv. phaseolicola (the "kudzu strain") and P.syringae pv. glycinea, which were efficient ethylene producers, several strains of P.syringae pvs. sesami and cannabina generated PCR products of the predicted size. A DNA probe of the efe gene, isolated from strain PK2, hybridized to these PCR products, indicating homology to the P.syringae pv. phaseolicola efe gene. PCR restriction fragment length polymorphism analyses suggested that these four pathovars harbor a similar efe gene. Furthermore, the probe hybridized to an indigenous plasmid of P.syringae pv. cannabina, suggesting that the efe gene could be located on a plasmid in this pathovar, but did not hybridize to plas-mids of P.syringae pv. sesami strains. P.syringae pvs. sesami and cannabina strains produced ethylene in King's medium B at levels similar to those of P.syringae pvs. phaseolicola and glycinea. Thus, two new ethylene-producing bacteria were detected by the PCR assay.  相似文献   

15.
ABSTRACT Strains of Pantoea agglomerans (synanamorph Erwinia herbicola) suppressed the development of basal kernel blight of barley, caused by Pseudomonas syringae pv. syringae, when applied to heads prior to the Pseudomonas syringae pv. syringae infection window at the soft dough stage of kernel development. Field experiments in 1994 and 1995 revealed 45 to 74% kernel blight disease reduction, whereas glasshouse studies resulted in 50 to 100% disease control depending on the isolate used and barley cultivar screened. The efficacy of biocontrol strains was affected by time and rate of application. Percentage of kernels infected decreased significantly when P. agglomerans was applied before pathogen inoculation, but not when coinoculated. A single P. agglomerans application 3 days prior to the pathogen inoculation was sufficient to provide control since populations of about 10(7) CFU per kernel were established consistently, while Pseudomonas syringae pv. syringae populations dropped 100-fold to 2.0 x 10(4) CFU per kernel. An application to the flag leaf at EC 49 (before heading) also reduced kernel infection percentages significantly. Basal blight decreased with increasing concentrations (10(3) to 10(7) CFU/ml) of P. agglomerans, with 10(7) CFU/ml providing the best control. For long-term preservation and marketability, the survival of bacterial antagonists in several wettable powder formulations was tested. Over all formulations tested, the survival declined between 10- to >100-fold over a period of 1.5 years (r = -0.7; P = 0.000). Although not significant, storage of most formulations at 4 degrees C was better for viability (90 to 93% survival) than was storage at 22 degrees C (73 to 79%). However, long-term preservation had no adverse effect on biocontrol efficacy.  相似文献   

16.
ABSTRACT Strains of Pseudomonas syringae pv. pisi and Ralstonia solanacearum produced ethylene at rates 20- and 200-fold lower, respectively, than strains of P. syringae pvs. cannabina, glycinea, phaseolicola, and sesami. In the current study, we investigated which ethylene biosynthetic pathways were used by P. syringae pv. pisi and R. solanacearum. Neither the activity of an ethylene-forming enzyme nor a corresponding efe gene homolog could be detected in R. solanacearum, suggesting synthesis of ethylene via 2-keto-4-methyl-thiobutyric acid. In contrast, 2-oxoglutarate-dependent ethylene formation was observed with P. syringae pv. pisi, and Southern blot hybridization revealed the presence of an efe homolog in this pathovar. The efe genes from P. syringae pvs. cannabina, glycinea, phaseolicola, pisi, and sesami were sequenced. Nucleotide sequence comparisons indicated that the efe gene in pv. pisi was not as highly conserved as it was in other P. syringae pathovars. The pv. pisi efe homolog showed numerous nucleotide substitutions and a deletion of 13 amino acids at the C-terminus of the predicted gene product. These sequence alterations might account for the lower rate of ethylene production by this pathovar. All ethylene-producing P. syringae pathovars were virulent on bush bean plants. The overlapping host range of these pathovars suggests that horizontal transfer of the efe gene may have occurred among bacteria inhabiting the same host.  相似文献   

17.
ABSTRACT From a collection of 152 pseudomonads isolated from diseased beans in Spain, 138 (91%) of the strains were identified as Pseudomonas syringae pv. phaseolicola and the rest as P. syringae pv. syringae. The P. syringae pv. phaseolicola strains produced typical water-soaked lesions on bean pods, although 95 of them did not produce phaseolotoxin in vitro. Ninety-four of these isolates did not produce the expected 0.5-kb product after polymerase chain reaction (PCR) amplification using primers specific for open reading frame (ORF) 6 of the phaseolotoxin (tox) gene cluster and did not contain DNA homologous to ORF 6 in Southern hybridization experiments. To our knowledge, this is the first report of the widespread occurrence in the field of strains of P. syringae pv. phaseolicola lacking the tox cluster, which contrasts sharply with the general belief that Tox(+) isolates are the only ones with epidemiological importance. Additionally, the tox(-) isolates were not specifically detected by a commercial polyclonal antisera in an enzyme-linked immunosorbent assay. Accordingly, it is possible that the certification of seed lots as free of the pathogen cannot be reliably done in Spain, or in any other country where tox(-) strains might occur frequently, using current PCR or serological protocols. The amplification of three avirulence genes by PCR allowed us to make predictions of the P. syringae pv. phaseolicola race structure, as confirmed by plant assays. Six races (races 1, 2, 5, 6, 7, and 9) were identified, with race 7 being the most prevalent (46.1%) followed by races 6 (21.3%) and 1 (9.0%). All the tox(-) isolates contained gene avrPphF, typical of races 1, 5, 7, and 9.  相似文献   

18.
猕猴桃品种酚类物质及可溶性蛋白含量与抗溃疡病的关系   总被引:4,自引:0,他引:4  
以安徽省猕猴桃主栽品种金魁、早鲜、魁蜜、华美2号、秦美、金丰为研究对象,于展叶孕蕾期分别取发病的枝条、叶片,以未发病健株的相应组织为对照,分析枝条、叶片中酚类物质和可溶性蛋白的含量变化。结果表明:抗病品种健株枝条、叶片中可溶性蛋白含量显著高于易感病品种,说明枝条中可溶性蛋白含量与品种抗性成正相关。自然发病后,感病品种枝条中可溶性蛋白含量增加,抗病品种可溶性蛋白含量降低。抗病品种健枝条、叶片中酚类物质含量高于易感病品种的健枝、叶,发病后抗感品种酚类物质含量都增加。  相似文献   

19.
Twenty-eight strains of Pseudomonas syringae pv. actinidiae isolated in 1984, 1987 and 1988 from kiwifruit orchards in Japan were tested for their resistance to copper sulfate. All strains isolated in 1984 were copper sensitive with a minimum inhibitory concentration (MIC) of cupric sulfate of 0.75 mM. However, some strains isolated in 1987 and 1988 were resistant, with the MIC ranging from 2.25 to 3.0 mM. All copper-resistant strains contained at least one of two plasmids, pPaCul (about 70.5 kb) or pPaCu2 (about 280 kb), or both. In a copper-resistant strain Pa429, the location of the copper-resistance gene(s) was examined by insertional inactivation with Tn5. The MIC of copper sulfate in the copper-sensitive mutant obtained by Tn5 tagging decreased from 2.75 to 0.75 mM. The 14.5 kb BamHI fragment, designated pPaCuB14, containing the same locus mutagenized with Tn5 was cloned from pPaCu1. However, pPaCuB14 did not confer copper resistance in the transformant of copper-sensitive strain Pa21R, suggesting that this clone did not contain a full set of copper-resistance gene(s). Then a cosmid library of pPaCu1 was constructed and six cosmid clones hybridized with pPaCuB14 were selected. One of the six cosmids, designated pPaCuC1, conferred a near wild-type level of copper resistance in the transformant of the copper-sensitive strain. pPaCuC1 had a homologous region that hybridized with all of the PCR-amplifled fragments of copA, copB, copR, and copS genes of P. syringae pv. tomato. DNA sequence analysis of the homologous region revealed the existence of four open reading frames (ORF A, B, R and S) oriented in the same direction. The predicted amino acid sequences of ORF A, B, R and S had 80, 70, 97 and 95% identity with CopA, B, R and S of P. syringae pv. tomato, respectively. Received 5 July 2001/ Accepted in revised form 27 September 2001  相似文献   

20.
A total of 101 Pseudomonas syringae pv. syringae strains, obtained from international culture collections or isolated from diseased tissues of herbaceous and woody plant species, were assessed by repetitive PCR using the BOX primer, and for the presence of the syrB gene. Representative strains were also tested for pathogenicity to lilac, pear, peach, corn and bean, as well as for virulence to lemon and zucchini fruits. The unweighted pair-group method using arithmethic averages analysis (UPGMA) of genomic fingerprints revealed 17 different patterns which grouped into three major clusters, A, B and C. Most of the strains (52·4%) were included in patterns 1–4 of group A. These patterns comprised strains obtained from either herbaceous or woody species, and showed four fragments of similar mobility. Genetic variability was ascertained for strains isolated from apple, pear, apricot, Citrus spp. and cereals. No clear relationship was observed between host plant and bacterial genomic fingerprint. Variability was also observed in pathogenicity and virulence tests. The inoculation of pear leaves discriminated strains isolated from pear as well as the very aggressive strains, whereas inoculation of lilac, peach and corn did not discriminate the host plant from which the strains were originally isolated. Lemon fruit inoculation proved very effective for P. syringae pv. syringae virulence assessment. The syrB gene was present in almost all strains.  相似文献   

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