Jae-Sung Jung

Comparative Analysis of Korean and Japanese Strains of Pseudomonas syringae pv. actinidiae Causing Bacterial Canker of Kiwifruit

Genomic and phenotypic characteristics of the bacterial strains of Pseudomonas syringae pv. actinidiae and P. syringae pv. syringae collected from several kiwifruit orchards of Korea were investigated and compared with those from Japan to elucidate their phylogenic relationships. All the strains of P. syringae pv. actinidiae and pv. syringae tested were sensitive to copper sulfate but Korean and Japanese strains showed quite different responses to streptomycin. Korean strains were sensitive to streptomycin, but most of the Japanese strains of P. syringae pv. actinidiae were highly resistant to streptomycin. Japanese strains were also relatively more resistant to oxytetracycline than Korean strains. Plasmid profiles were not valuable to distinguish Korean strains of P. syringae pv. actinidiae frombJapanese strains. One or more indigenous plasmids with more than 15 kb in size were detected in all strains of P. syringae pv. actinidiae, but the number and sizes of plasmids harbored in P. syringae pv. actinidiae were variable among the strains regardless of their geographic origins. There also observed no significant relationship among resistance levels of the strains of P. syringae pv. actinidiae to antibiotics, their pathogenicity and plasmid profiles. RAPD profiles were useful to analyze the strains of P. syringae pv. actinidiae and pv. syringae. All the strains of P. syringae pv. actinidiae fell into a wide cluster separated from the strains of P. syringae pv. syringae, but Korean strains of P. syringae pv. actinidiae were separated from Japanese strains. The results support that Korean and Japanese strains of P. syringae pv. actinidiae may have different phylogenic origins.

Optimum Spray Program of Preventive Fungicides for the Control of Postharvest Fruit Rots of Kiwifruit

Fungicides of tebuconazole wp, iprodione wp and flusilazole wp were applied for the control of postharvest fruit rots of kiwifruit (Actinidia deliciosa) in the field in 2000 and 2001. More than 3 consecutive applications of these fungicides from the late June with 10-day-interval successfully controlled the diseases. It was found in the field trial in 2002 that 4 consecutive spays from mid of June with 10-day-interval was found to be the most effective application program for tebuconazole wp, iprodione wp and flusilazole wp, The results suggested that currently registered fungicides of benomyl wp and thiophanate-methyl wp can be substituted by tebuconazole wp, iprodione wp and flusilazole wp for the control of the diseases in Korea. Use of these fungicides can restrain emergence of fungicide resistant strains of postharvest fruit rot pathogens with benefit of reduced application of chemicals for food safety and environmental conservation.

Causal Agents of Blossom Blight of Kiwifruit in Korea

The causal agents of bacterial blossom blight in kiwifruit were isolated from flowers displaying symptoms in Korea. The pathogens were characterized by biochemical and physiological tests, and identified on the basis of 16S rDNA and 16S-23S internal transcribed spacer (ITS) sequences. Pathogenicity tests demonstrated that the blossom blight of kiwifruit in Korea is caused by two pathogens, Pseudomonas syringae pv. syringae and P. fluorescens. Carbon source utilization and DNA-DNA hybridization experiments confirmed P. fluorescens as one of the causal agents of blossom blight of kiwifruit. P. syringae pv. syringae and P. fluorescens can be distinguished from each other by the symptoms they produce in flowers. P. syringae pv. syringae primarily affected the stamen, while P. fluorescens caused rotting of all internal tissues of buds or flowers.

Effect of Ripening Temperatures on Incidences of Postharvest Fruit Rots of Kiwifruits

This study was conducted to identify optimum ripening condition for kiwifruits (Actinidia deliciosa) to prevent postharvest fruit rots caused by Botryosphaeria dothidea, Diaporthe actinidiae and Botrytis cinerea. The optimum temperatures for mycelial growth of B. dothidea, D. actinidiae and B. cinerea were , and , respectively, and the incidence was closely related with the temperature. Although kiwifruits ripened faster at higher temperatures, the rates of diseased fruits increased with the rates of ripened fruits increased. Optimum conditions for ripening of kiwifruit were 20-day at 17C.

Incidences and Causal Agents of Postharvest Fruit Rots in Kiwifruits in Korea

Sixteen hundred fruits were randomly collected from 16 kiwifruit (Actinidia deliciosa) orchards in Jeonnam, Gyeongnam and Jejn provinces in Korea in 2000 and incidences of postharvest fruit rots were examined. The overall disease incidence was 32% and varied much with locations of orchards ranging from 5 to 68%. The percentages of kiwifruits showing internal, external, and both internal and external symptoms were 21.9%, 4.9%, and 5.2%, respectively. Several fungi were isolated from rotten fruits; Botryosphaeria dothidea, Diaporthe actinidiae and Botrytis cinerea were the major pathogens with the average isolation rates of 83.3%, 11.9% and 1.4%, respectively. Based on the symptoms on kiwifruits and the characteristics, the postharvest fruit rots caused by B. dothidea and D. actinidiae are suggested to be named as ripe rots and stem-end rots, respectively.

Identification and Ecological Characteristics of Bacterial Blossom Blight Pathogen of Kiwifruit

Bacterial blossom blight is one of the most important diseases of kiwifruit (Actinidia deliciosa). The disease occurs during flowering in the late May and disease outbreaks associated with rainfall during the flowering season have resulted in a severe reduction in kiwifruit production. The causal organism isolated from diseased blossoms of kiwifruits was identified as Pseudomonas syringae pv, syringae based on the physiological and biochemical characteristics and pathogenicity test. Dead fruit stalks, dead pruned twigs, fallen leaves and soils mainly provided R syringae pv. syringae with overwintering places in the kiwifruit orchards, and the inocula also overwintered on buds, trunks, branches, and twigs on the kiwifruit trees. Among the overwintering places, the incula were detected in the highest frequencies from dead fruit stalks. The population density of P. syringae pv. syringae was speculated to be over cfu/ml for the bacterial infection, and the optimum temperature for the bacterial growth ranged 20 to . The highest population density of P. syringae pv. syringae on the overwintering places was detected in May and June when the daily average temperature coincided with the optimum temperature for bacterial growth of P. syringae pv. syringae.

Nested PCR Detection of Pseudomonas syringae pv. actinidiae, the Causal Bacterium

A PCR method that combines biological and enzymatic amplification of PCR targets was developed for the detection of Pseudomonas syringae pv. actinidiae on kiwifruit leaves. A nested PCR was performed with primers designes from the coding sequence of the cfl gene, which is involved in production of the phytotoxin coronatine. The first and second primer sets efficiently amplified expected 665 and 310-bp fragments, respectively. With two successive amplifications, as few as 20 CFU/ml of P. syringae pv. actinidiae could be detected on ethidium bromide-stained agarose gel. Leaf samples were collected from 4 kiwifruit trees showing yellow halo spots on leaves and incubated in pepton-sucrose broth for 12 h at C before PCR amplification. Positive detection was obtained with one sample, which was proved as a diseased plant in the next spring.

Functional characterization of the sucrose isomerase responsible for trehalulose production in plant-associated Pectobacterium species

Fifty-three plant-associated microorganisms were investigated for their ability to convert sucrose to its isomers. These microorganisms included one Dickeya zeae isolate and 7 Enterobacter, 3 Pantoea, and 43 Pectobacterium species. Eleven out of the 53 strains (21%) showed the ability to transform sucrose to isomaltulose and trehalulose. Among those, Pectobacterium carotovorum KKH 3-1 showed the highest bioconversion yield (97.4%) from sucrose to its isomers. In this strain, the addition of up to 14% sucrose in the medium enhanced sucrose isomerase (SIase) production. The SIase activity at 14% sucrose (47.6 U/mg dcw) was about 3.6-fold higher than that of the negative control (13.3 U/mg dcw at 0% sucrose). The gene encoding SIase, which is comprised a 1776 bp open reading frame (ORF) encoding 591 amino acids, was cloned from P. carotovorum KKH 3-1 and expressed in Escherichia coli. The recombinant SIase (PCSI) was shown to have optimum activity at pH 6.0 and 40 °C. The reaction temperature significantly affected the ratio of sucrose isomers produced by PCSI. The amount of trehalulose increased from 47.5% to 79.1% as temperature was lowered from 50 °C to 30 °C, implying that SIase activity can be controlled by reaction temperature.

Streptomycin Resistant Genes of Pseudomonas syringae pv. syringae, the Causal Agent of Bacterial Blossom Blight of Kiwifruit

A total of 41 Pseudomonas syringae pv. syringae, the causal agent of bacterial blossom blight, were isolated from kiwifruit plants in Korea. Among them, two strains showing streptomycin resistance were examined to investigate the structure of resistant determinants by PCR and nucleotide sequence analysis. PCR results suggested that the streptomycin resistance is mediated by strA-strB genes carried on Tn5393a. Insertion sequences, IS6100 and IS1133, which were located within or downstream of tnpR gene in Xanthomonas campestris and Erwinia amylovora were not found. Nucleotide sequences of strA-strB were 100% identical with Tn5393a. Two stretomycin resistant strains had three plasmids. Southern blot hybridization using strA-strB probe indicated that the resistant genes were carried on a 100kb plasmid.

Optimum Spray Program of Preventive Bactericides for the Control of Bacterial Blossom Blight of Kiwifruit

Bacterial blossom blight of kiwifruit (Actinidia deliciosa) has been mainly controlled by antibiotics. Among 15 candidate chemicals, streptomycin sulfate oxytetracycline WP, streptomycin copper hydroxide WP and oxolinic acid WP were selected as preventive bactericides against bacterial blossom blight of kiwfruit through in vitro and in vivo test. Spray of streptomycin sulfate oxytetracycline WP and streptomycin copper hydroxide WP at flowering period was most effective in controlling bacterial blossom blight of kiwifruit. Among the various combinations of spray times at different spray periods, optimum spray program of the preventive bactericides for the control of bacterial blossom blight of kiwifruit was turned out to be 3 times application with 10 day-interval from early May during the flowering season of kiwifruits.