Abstract of Presentations

Abstract of Presentations

266

Abstract of Presentations Abstracts presented at the 2009 KSPP Annual Fall Meeting and the 1st Japan-Korea Joint Symposium of the Korean Society of Plant Pathology in Jeju KAL Hotel, Jeju, Korea, October 28-31, 2009.

A-1

Effect of simulated acid rain on the development of pepper anthracnose. Sun Mi Kang, Yuliang Gao and Byeongjin Cha*. Department of Plant Medicine, Chungbuk National University, Cheongju 361-763, Korea

Changes in the development of an anthracnose disease (Colletotrichum acutatum) according to simulated acid rain (SAR) treatment to pepper fruits and seedlings were investigated. The pH levels of SAR used in the experiment were 6.5(control), 5, 4, 3, and 2. In laboratory tests, mycelial growth was suppressed by the decrease in pH of a culture medium (PDA): mycelial growth of C. acutatum on PDA of pH4 was suppressed 15% compared to control. Also, low pH suppressed spore germination of C. acutatum to a great extent: spore germinations rate was 0.1% and 2%, respectively in pH2 and pH3, although the rate was 96% in control. In in vivo tests with pepper fruits (cv. Nokgwang), longer dipping-time to SAR (sulfuric acid: nitric acid=2:1) resulted higher disease severity: the highest disease severity was recorded in the treatment 60 minutes dipping to SAR among 20, 40 and 60 minutes dipping. In greenhouse tests, pepper seedlings (cv. Hanyeoreum bigarim) were planted into the mixture of gardening soil and field soil (1:1, pH5.8). When the seedlings grown to 6-8 leafstage, seedling pots were moved to the SAR dripping equipment and treated with SAR about 30 minutes/time, 3 times/week, for 2 to 3 weeks. Disease severity was higher in the treatment of pH3, pH4 compared to the control. However, seedlings of control showed higher disease severity than seedlings in pH2. The visible damage was appeared on pepper seedlings, mostly on the leaves, in pH2-treatment as a white and irregular spots with pale brown margin. Those white lesions were expanded and the numbers were increased with repeated treatment of SAR. Affected leaves wilted and defoliated easily.

B-1

Diversity and genetic characterization of thymidylate kinase gene of phytoplasma associated with jujube witches’-broom phytoplasma in China. Qi-cong Xu , Guo-zhong Tian , Cai-li Lin , Yong 1,2

1

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Li , Chun-gen Piao and Hui-lian Xu . Research Institute of Forest 1

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Ecology, Environment and Protection, Chinese Academy of Forestry, Beijing 100091, China ; International Nature Farming Research Center, 5632 Hata, Nagano 390-1401, Japan

Thymidylate kinase (TMK, EC 2.7.4.9), which catalyses the transfer of a terminal phosphoryl group from ATP to dTMP, is crucial to both the de novo synthetic and the salvage pathways for pyrimidine deoxyribonucleotides. Three types of thymidylate kinase genes, tmk-x, tmk-y and tmk-z (TMK gene), of the jujube (Zizyphus jujube Mill) witches-broom (JWB) phytoplasma were identified by means of sequence analyses as well as RFLP and SSCP of the PCR products in

34 JWB infected jujube samples in China. Tmk-x and tmk-y were identified in many jujube cultivars from different regions, whereas tmk-z was only found in BJLZ strain infecting an aged dragon jujube from Beijing. Tmk-x and tmk-y shared the similarity of 92.33 and 90.90% with the tmk-a gene (AB010446) in OY-W strain of onion yellows, whereas tmk-z shared 97.89% similarity with one of tmk gene copies in genome of OY-M (AP006628, from 391588 to 392202) of onion yellows. All three deduced proteins, TMKx, TMKy and TMKz of these tmk genes contained conserved motifs for catalytic activity. The detected JWB samples from different cultivars with various geographic origins and different resistance to the JWB could be mainly classified into three types, typeX, typeY and typeXY, containing tmk-x gene, tmk-y gene and both tmk-x and tmk-y genes, respectively. Strains of typeX and typeY seemed to be detected mainly in susceptible cultivars, while typeXY was likely to be much related to resistant cultivars. It is assumed that the formation of distinct tmk genotypes among JWB strain populations might be associated with the phytoplasma genetic variation related to the cultivar resistance and environment as well as jujube germplasm exchange.

B-2

First report of Acidovorax avenae subsp. avenae on Millet (Panicum miliaceum) in Korea. Young Nam Yoon , Young Kee Lee , Ji 1

2

Hun Jung , Yeong Hoon Lee , Hyun Joo Kim , Soon Do Bae , Byeong Ryeol Choi and Min Hee Nam . Department of Functional 1

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Crop, NICS, RDA, Miryang, 627-803, Korea; Department of Agricultural Biology, NAAS, RDA, Suwon, 441-857, Korea 2

Diseased millet(Panicum miliaceum) which were collected in Bonghwa, Miryang region and other places in Korea, showed brown stripes on leaf sheaths. They were Gram negative. Results using Biolog GN MicroPlates (Biolog MicroLog 3 Release 4.20) showed all strains to be A. avenae subsp. avenae. Hypersensitive reactions on leaves of 1-month-old tobacco plants infiltrated with bacterial suspensions were recorded. Strains were tested for pathogenicity by injecting stems of 20-day-old millet plants with bacterial suspensions (approximately 10 CFU/ml). Inoculated millet was incubated for 5 days under humid conditions at 28°C. Inoculated millet plants showed brown stripes and non-inoculated control millet remained symptomless. Based on biochemical, serological, and biological characteristics, strains were identified as A. avenae subsp. avenae. This is the first report of A. avenae subsp. avenae, causal agent of bacterial stripe of the millet in Korea. 8

B-3

Role of Siderophore pyoverdine in the virulence of Pseudomonas syringae pv. tabaci 6605. Yuki Ichinose , Tomoko Suzuki , Yoshish1

1

ige Inagaki , Kazuhiro Toyoda , Tomonori Shiraishi and Fumiko 1

1

1

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Taguchi . The Graduate School of Natural Science and Technology, 1

1

Okayama University, Okayama 700-8530, Japan

To investigate the role of siderophore pyoverdine- mediated iron uptake in the virulence of Pseudomonas syringae pv. tabaci 6605 (Pta6605), three predicted pyoverdine synthesis related genes, pvdJ, pvdL and fpvA, were mutated. The genes pvdJ, pvdL and fpvA encoded the pyoverdine side-chain peptide synthetase III L-Thr-L-Ser component, pyoverdine chromophore and TonB-dependent ferripyoverdine receptor, respectively. The ∆pvdJ and ∆pvdL mutants were unable to produce pyoverdine in MG medium as the iron-depleted condition. Furthermore, the ∆pvdJ and ∆pvdL mutants showed lower abilities to produce tabtoxin and acyl homoserine lactones (AHL), quorum-sensing molecules, and consequently had reduced virulence on host tobacco plants. In contrast, all of the mutants had accelerated swarming ability and increased biosurfactant production, suggesting that swarming motility and biosurfactant production might be negatively controlled by pyoverdine-mediated iron uptake. Scanning electron micrographs of the surface of tobacco leaves inoculated with the mutant strains revealed only small amounts of extracellular polymeric matrix around these mutants, indicating disruption of mature biofilm. The tolerance to antibiotics was drastically increased in the ∆pvdL mutant, similar to the ∆psyI mutant, which is defective in AHL production. These results demonstrated that pyoverdine synthesis and the quorum sensing system of Pta6605 are indispensable for virulence in host tobacco infection, and AHL may negatively regulate the tolerance to antibiotics.

B-4

Role of flagellar motility in the virulence of Pseudomonas syringae pv. tabaci. Eiko Kanda , Fumiko Taguchi , Tomoko Suzuki , 1

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Yoshishige Inagaki , Kazuhiro Toyoda , Tomonori Shiraishi and Yuki Ichinose . The Graduate School of Natural Science and Tech1

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1

nology, Okayama University, Okayama 700-8530, Japan

Pseudomonas syringae pv. tabaci 6605 (Pta6605) is a causal agent for

wildfire disease in tobacco. Recently, we found that flagellin, a major constituent in the flagella filament of this pathogen, is a potent elicitor of hypersensitive reaction in non-host plant species. We also found that flagellin is required for virulence against the host plant. In this study, we investigated the role of flagellar motility in bacterial virulence. We introduced mutation into fliC, motAB and motCD and generated motility-defective mutant strains. The fliC encodes flagellin protein, whereas each set of motAB and motCD encodes stator part of motor complex. The mutant strains ∆filC lost motility and reduced the virulence. The mutant strain ∆motCD also lost motility, but ∆motAB retained motility. Therefore MotCD seems to be major components of motor stator. Interestingly all flagellar motility-defective mutant strains, ∆fliC and ∆motCD also lost the ability to produce acylhomoserine lactones (AHL). Thus flagellar motility and AHL production seems to be related. To confirm the relationship between motility and ability to produce AHL, we used flagellin-glycosylation defective mutant strains. Flagellin from Pta6605 is glycosylated at 6 serine residues, and the mutant strains in which glycosylated Ser was substituted to Ala were partially motility-defective. Thus we found that the positive relationship between motility and AHL production in Pta6605. Furthermore, we found that orf3 gene which encodes 3oxoacyl-(acyl carrier protein) synthase III which is required for AHL

production upstream of fliC in flagella gene cluster. These results indicate that flagella motility is strongly related to the ability to produce AHL in Pta6605.

B-5

Genetic analysis of genes involved in synthesis of modified 4amino-4,6-dideoxyglucose in flagellin of Pseudomonas syringae pv. tabaci. Nguyen Linh Chi , Fumiko Taguchi , Masanobu 1

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Yamamoto , Mayumi Ohnishi-Kameyama , Mitsuru Yoshida , Tadashi Ishii , Tomoyuki Konishi , Kazuhiko Tsunemiv and Yuki Ichinose . The Graduate School of Natural Science and Technology, 2

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Okayama University, Okayama 700-8530, Japan. National Food Research Institute, Ibaraki 305-8642, Japan. Forestry and Forest Products Research Institute, Ibaraki 305-8687, Japan 2

3

Glycosylation of flagellin is essential for bacterial virulence and seems to be also involved in the determination of host specificity. Flagellins from Pseudomonas syringae pv. tabaci 6605 has been found to be glycosylated at six serine residues (position 143, 164, 176, 183, 193 and 201). The glycan structure at serine 201 was determined to consist of two L-rhamnoses and one modified 4-amino-4,6dideoxyglucose (viosamine). Furthermore, other serine residues have been expected to possess the similar trisaccharide units. To clarify the mechanisms for glycosylation of modified viosamine, genes encoding dTDP-viosamine aminotransferase (vioA), dTDP-viosamine acetyltransferase (vioB), and viosamine-derivative transferase (vioT) were isolated and their defective mutants were generated. Both immunoblot and MALDI-TOF MS analyses of flagellin showed that flagellin molecular masses of three mutants were reduced compared to that of wild-type. MALDI-TOF MS analysis of a lysyl endopeptidase-digested peptide including all six glycosylation sites from each flagellin indicated that the glycopeptides obtained from mutants had glycans consisting only of deoxyhexose instead of the flagellin glycans including the viosamine derivatives determined previously. In addition, the motility and virulence on host tobacco leaves were strongly impaired in ∆vioA mutant and were weakly reduced in ∆vioB and ∆vioT mutant strains. In Escherichia coli VM187 strain (O7:K1), N-acetylviosamine was found as a component of O7-specific lipopolysaccharide (LPS), and vioA was identified to be required for the synthesis of O7-specific LPS. However, in this study, the banding patters of LPS were similarly observed by SDS-PAGE analysis in both P. syringae pv. tabaci 6605 wild-type and three mutants. These results suggest that the genes vioA, vioB, and vioT are specific and essential for glycosylation of flagellin, and accordingly are required for bacterial virulence.

B-6

The aldehyde dehydrogenase gene and phosphinothricin Nacetyltransferase gene located in pathogenicity island is required for virulence of Pseudomonas cichorii on eggplant but not on lettuce. M. Tanaka, K. Ohnishi, A. Kiba and Y. Hikichi. Laboratory of Plant Pathology & Biotechnology, Kochi University, Nankoku, Kochi 783-8502, Japan

Pseudomonas cichorii strain SPC9018 (SPC9018) causes necrotic

lesion and rot on eggplant and lettuce, respectively. The nucleotide sequences of pathogenicity island (PAI) including the hrp genes of

Abstract of Presentations SPC9018 and its genetic structure are homologous to those of S-PAI of P. viridiflava BS group strains. The consensus hrp-box (GGAACCN -CCANNCA) was identified at putative promoters of hrpA, hrpF, hrpW, avrE, avrF and hrpJ. RT-PCR analysis showed that expression of hrp genes was dependently on HrpL. Interestingly, pathogenicity of SPC9018 on eggplant are hrp-dependent but not on lettuce (Hojo et al. 2008). In PAI of SPC9018, LysR-type transcription regulator gene (LysR), aldehyde dehydrogenase gene (aldh) and phosphinothricin N-acetyltransferase gene (pna) are located adjacent to hrpL. RTPCR analysis showed that expression of aldh and pna is independent of HrpL. The aldh-deficient mutant (∆aldh) and pna-deficient mutant (∆pna) of SPC9018 lost their virulence on eggplant, but retained their virulence on lettuce. Whereas LysR-deficient mutant retained their virulence on both eggplant and lettuce. Complementing the ∆aldh and ∆pna with aldh and pna originating from SPC9018, respectively, allowed the mutants to cause the disease on eggplant. Furthermore, aldh and pna from S-PAI of P. viridiflava strain Pv9508 (BS group) also allowed the mutants to complement their virulence on eggplant. These results suggest that requirement of aldh and pna located in PAI for virulence of SPC9018 on eggplant and functional conservation of those genes between P. cichorii and P. viridiflava. 15-16

B-7

Regulation of universal stress proteins by quorum sensing in Burkholderia glumae. Hongsup Kim, Yongsung Kang, Eunhye Goo and Ingyu Hwang. Department of Agricultural Biotechnology, Seoul National University, Seoul, 151-921, Korea

Burkholderia glumae has a TofI/TofR quorum sensing system that enables cell to cell communication using of N-octanoyl homoserine lactone (C8-HSL) as a signal. TofI of B. glumae is the autoinducer

synthase responsible for C8-HSL synthesis, and TofR is a cognate receptor. TofR/C8-HSL regulates toxoflavin production and expression of qsmR, which is an IclR type transcriptional regulator. We used the two-dimensional gel electrophoresis and mass spectrometry techniques to identify proteins of B. glumae important for pathogenicity and quormone responses. Proteome analysis revealed that C8-HSL produced by B. glumae significantly induced or reduced the level of different proteins. Among those, universal stress protein (UspA) encompassing a conserved group of proteins that are found in bacteria, archaea and eukaryotes was of interest. In E. coli, UspA proteins are general responders to stresses and present one of the most abundant proteins in stationary phase cells. B. glumae has eleven uspA genes similar to uspA in E. coli. We demonstrate that expression of uspA1 and uspA2 of B. glumae is regulated by QsmR. To determine the functions of UspA1 and UspA2, we evaluated how the uspA1 and uspA2 mutants survive under many different types of stress. We found that survival of the mutants was decreased dramatically after two hours incubation at 45 C with rigorous shaking. This indicated that UspA1 and UspA2 are essential for the survival of the bacterium during heat shock. This is the first report that the uspA gene expression is regulated by quorum sensing. o

B-8

Sigma S as a Regulator for Early Infection and Bacterial Survival in Ralstonia solanacearum. Hye Kyung Choi , Eun Jin Park , Hae 1

2

Young Um , Byung Ju Moon and Seon-Woo Lee . Department of 1

2

1,2

1

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Medical Bioscience, Dong-A University, Busan 604-714, Korea; Department of Applied Biology, Dong-A University, Busan 604-714, Korea 2

Ralstonia solanacearum causes lethal wilt of the solanaceous plants

and survives over seasons in natural soil ecosystem. Sigma S has been shown to regulate gene expression during stationary phase or in response to stress conditions in other bacteria. To investigate the role of sigma S in R. solanacearum under different stress conditions, we constructed a sigma S mutant of this bacterium. The rpoS gene encoding sigma S was cloned from R. solanacearum GMI1000, and then we performed marker exchange mutagenesis. Gene replacement in the mutant strain was confirmed by Southern hybridization. No differences in the colony morphology and extracellular polysaccharide production were observed upon comparison between rpoS mutant and the wild type strain. Both wild type and the mutant strain showed the same activity for pectin degradation, while the rpoS mutant exhibited reduced cellulase activity. RT-PCR analysis revealed that two endoglucanase gene expression were down-regulated in the rpoS mutant, compared to wild type. When stored in phosphate buffered saline solution and CPG broth at 4 C, the viability of the rpoS mutant decreased rapidly over time in comparison to the wild type strain. Furthermore, the virulence of the mutant decreased significantly upon infection of the tomato plant. Our results from rpoS mutant suggested that sigma S might be a regulator for early stage of bacterial infection to tomato plants. Under stress conditions, wild type strain was more sensitive to starvation but less sensitive to hydrogen peroxide exposure than rpoS mutant. The role of rpoS gene in R. solanacearum was further confirmed through rpoS gene complementation in trans by restoring the original phenotypes. The long term survival of the wild type strain and the rpoS mutant is being examined in artificial soil microcosm system at different environmental conditions. o

B-9

A Chromosomal Locus Involved in Extracellular Polysaccharide Production in Ralstonia solanacearum SL341. Hae Young

Um , Hye Kyung Choi , Byung Ju Moon , and Seon-Woo Lee . 1

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2

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Department of Medical Bioscience, Dong-A University, Busan 604-714, Korea; Deparment of Applied Biology, Dong-A University, Busan 604-714, Korea 1

2

Ralstonia solanacearum is the causal agent of bacterial wilt of numerous economically important crops worldwide. R. solanacearum pro-

duces large amounts of the exopolysaccharide I (EPS), which is a major virulence factor for efficient bacterial colonization in plant xylem to cause plant wilt. To investigate role of epsD related to extracellular polysaccharide production, we constructed epsD defective mutant from R. solanacearum SL341 strain by insertion of kanamycin resistant gene. We obtained three EPS-nonproducing (eps-) mutants which had nonmucoid colony morphology. However, Southern blot analysis revealed that all three mutants were intact in target epsD gene but they showed the gene replacement in the other chromosomal locus. Analysis of the nucleotide sequence of disrupted chromosomal locus of three mutants revealed approximately 3 kb DNA deletion encoding zonula occludens toxin homologues and bacteriophage hypothetical proteins. However the chromosomal locus was not present in the genome sequences of other R. solanacearum strains such as GMI1000 and MolK2. Wild type and eps- mutant of R.

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solanacearum were compared for their virulence and hypersensitivity

on various plants, EPS production, copper sensitivity and survival in soils. The virulence of mutants certainly disappeared upon infection of tomato plants and Arabidopsis plant. Nevertheless, mutants showed hypersensitive response on tobacco. No differences in viability under stress condition between the mutant and the wild type were observed. To determine the role of deleted chromosomal locus from the eps- mutant, additional genetic analysis and complementation analysis are in progress.

B-10

A Flamentous Bacteriophage with Single Strand DNA Infecting Ralstonia solanacearum Strains. Ju-Young Bae , Senthilkumar 1

Murugaiyan , Seung Don Lee , Jing Wu , Nam Hee Kim and SeonWoo Lee . Department of Medical Bioscience, Dong-A University, 2

1,2

3

2

2

1

Busan 604-714, Korea; Department of Applied Biology, Dong-A University, Busan 604-714, Korea; Research Coordination Division, Rural Development Administration, Suwon 441-707, Korea 2

3

Ralstonia solanacearum is a soil-borne Gram-negative bacterium

which is a causative agent of bacterial wilt in many important crops. In this study, we isolated several bacteriophage from soil samples collected from different crop fields (tomato, hot pepper and tobacco) located in Korea. All the bacteriophages were shown to form clear plaques on CPG plate after 2 days of incubation with R. solanacearum. Out of the several bacteriophages, three bacteriophages were found to have a wide host range and were shown to infect eight different strains of R. solanacearum including two races, 1 and 3 as well as biovars 2, 3 and 4. Transmission electron microscopy revealed the presence of a filamentous structure in all the three bacteriophages. The purified phage DNAs were not affected by digestion with various restriction enzymes and RNase a but disappeared after nuclease S1 treatment, which specifically digests single stranded DNA (ssDNA). It was, therefore, concluded that the genomic DNAs of all three bacteriophages are single-stranded. We adopted multiple displacement amplification (MDA) to produce double stranded DNA from ssDNA of the bacteriophages. MDA products were randomly digested with restriction enzymes and cloned to analyze the genome sequences. DNA sequence analyes revealed that 9 open reading frames were present on the three bacteriophage, Genome analysis and electron microscopy indicated that three wide host range phages are members of Inoviridae. Due to the wide host range nature of three bacteriophages, they might be used as a biocontrol agent for bacterial wilt. Researches to use the wide host range phages for bacterial wilt control are in progress.

B-11

Race specific and rapid detection of Ralstonia solanacearum by direct PCR in the potato. Young-Gyu Lee , Jeom-Soon Kim and 1

1

Young-Kee Lee . Highland Agriculture Research Center, National Institute of Crop Science, Rural Development Administration, Pyeongchang 232-955, Korea; National Academy of Agricultural Science, Rural Development Administration, Suwon 441-707, Korea 2

1

2

Ralstonia solanacearum is the causative agent of bacterial wilt in

many important crops. Traditionally, the pathogen has been classified in five biovars according to carbon source utilization and in six

races based on host range. Major races of R. solanacearum in potato fields of Korea were into race 1 and 3. The aim of this study was to develop a race specific and rapid detection methods of R. solanacearum in the potato by direct PCR. Analysis of the 16s rRNA genes sequences of R. solanacearum was used to design a 13 primers for specific detection of the genus Ralstonia, R. solanacearum and R. solanacearum race 1 and 3. These primers were determined of compared to 68 published sequence of R. solanacearum isolates and the closely related 38 published sequence of the genus Ralstonia. To determine the specificity of these primers, direct PCRs were carried out with bacterial cell of all of the 23 isolates; R. solanacearum race 1 isolates(9), R. solanacearum race 3 isolates(5), the genus Ralstonia (3) and the others bacteria(6). In result, the 4 pairs of optimum direct-PCR primer combinations were selected. The PCR amplification using the sets I(RS68F + RS451RA) primers amplified a 412-bp DNA fragment for only R. solanacearum race 3 isolates tested. No amplification was observed with others isolates tested. The PCR amplification using the sets II(RS68F + RS451RB) primers amplified a 412-bp DNA fragment for only R. solanacearum race 1 isolates tested. The sets III(RS68F + R629R) primers amplified a 588-bp DNA fragment for R. solanacearum isolates tested. The sets IV(R97F + R908R) primers amplified a 834-bp DNA fragment for the genus Ralstonia isolates tested. The optimum primer combinations and concentrations and PCR annealing temperature and extension time were determined for the direct PCR. Direct PCRs were carried out with 7 samples showing wilt symptoms in the potato fields. The sample from potato plants were prepared with cutting of steams and dipping into sterilized distilled water for 20 minute. The specific band of R. solanacearum race 1 and 3 appeared in the all samples of naturally infected potato plants. The developed direct PCR methods for detecting the R. solanacearum races were very easy, rapid and powerful.

B-12

Detection of Pectobacterium carotovorum pv. actinidiae, a causal agent of a novel bacterial canker on kiwifruit ‘cv. Hort16A’ by PCR using primers based on DNA sequence of the carA gene.

Young Sun Lee , San Ho Son , Gyoung Hee Kim , Young Jin Koh and Jae Sung Jung . Department of Biology, Department of Plant 1

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Medicine, Sunchon National University, Suncheon 540-742, Korea

A causal agent of new bacterial canker characterized by die back or blight symptoms on young canes, often with red-rusty exudation on trunks during summer and autumn season was identified as Pectobacterium carotovorum subsp. actinidiae in previous work. A PCRbased method that utilizes primers to target the carA, carbamoylphosphate synthetase small subunit gene, was developed for the rapid and reliable detection of the pathogen. The PCR product amplified with primers specific to the carA gene of enterobacteria and genomic DNA of P. carotovorum subsp. actinidiae was cloned and the nucleotide sequence was determined. The sequence was aligned with those of the species belonging to genus Pectobacterium, which was obtained from GenBank database. On the basis of the sequence of the amplified carA region, a PCR primer set CarF/R specific to P. carotovorum subsp. actinidiae was designed. This primer set yielded a 343-bp product from DNA of P. carotovorum subsp. actinidiae, but no products from five other Pectobacterium spp. or from 11 epiphytic bacteria. Southern blot hybridization with the probe derived from the

Abstract of Presentations amplified product confirmed the PCR results. The Car primer system was successfully applied to detect the pathogen from the environmental samples.

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strain YC6903T and N. pyridinolyticus OS4 was 52.5%. The cellwall peptidoglycan type is LL-diaminopimelic acid. Based on the phenotypic, phylogenetic and chemotaxonomic data, the strain YC6903 represents a novel species of the genus Nocardioides, for which the name Nocardioides carexicola sp. nov. is proposed. The type strain is YC6903 (= KACC 13778 = DSM 22177 ). T

T

B-13

T

Sudden outbreak and rapid spread of Pseudomonas canker on the kiwifruit cv. ‘Hort16A (Actinidia chinensis Planchon)’ caused by Pseudomonas syringae pv. actinidiae in Korea. Gyoung Hee

Kim , Jae Sung Jung , Young Sun Lee , Jae-Seoun Hur and Young Jin Koh . Department of Plant Medicine, Department of Biology and 1

1

1

1

2

1

Department of Environmental Education, Sunchon National University, Suncheon 540-742, Korea 2

T

T

B-15

Analysis of the role of rpoE gene on virulence and stress response of Dickeya dadantii 3937. Md. Shahinur Kabir , Hisae Hirata and 1

2

Shinji Tsuyumu . Institute for Genetic Research and Biotechnology, 1,2

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836 Ohya, Suruga-ku, Shizuoka 422-8529, Shizuoka University, Japan; Faculty of Agriculture, 836 Ohya, Suruga-ku, Shizuoka 4228529, Shizuoka University, Japan Dickeya dadantii 3937 (formerly Erwinia chrysanthemi 3937) is the 2

Pseudomonas canker was first observed on the kiwifruit cv. ‘Hort16A (Actinidia chinensis)’ in Jeju Province, Korea in spring 2006. Die

back or blight on young canes, often with red-rusty exudation on canes or trunks and dark brown irregular spots surrounded with yellowish halos on leaves were its typical symptoms, which were closely resembled those on the kiwifruit cv. ‘Hayward (Actinidia deliciosa)’. Sudden outbreak and rapid spread of the Pseudomonas canker epidemics resulted in death of severely infected vines and eradication of completely devastated orchards of the kiwifruit cv. ‘Hort16A’. Pseudomonas canker has been observed at 11 kiwifruit cv. ‘Hort16A’ orchards since 2006, out of which four orchards were eradicated until now due to the severe epidemics. Contaminated pruning shears and climatic conditions seem to be responsible for sudden outbreak and rapid spread of the epidemics on all of the kiwifruit cv. ‘Hort16A’ vines in a short period. The causal bacterium was isolated from diseased vines of the kiwifruit cv. ‘Hort16A’ and identified as Pseudomonas syringae pv. actinidiae, the same Pseudomonas canker pathogen of the kiwifruit cv. ‘Hayward’ by morphological, cultural, physiological and biochemical characteristics and pathogenicity tests. Molecular detection of cfl gene encoding coronafacate ligase suggested that P. syringae pv. actinidiae strains isolated from the kiwifruit cv. ort16A as well as cv. Hayward in Korea produce coronatine instead of phaseolotoxin which was commonly produced by P. syringae pv. actinidiae strains isolated from the kiwifruit cv. Hayward in Japan.

B-14

A Novel Endophytic Bacterium, Nocardioides sp. YC6903 with Activities of Plant Growth Promotion and Salt Tolerance. Geun Cheol Song , Fehmida Bibi , Eu Jin Chung and Young Ryun Chung . Division of Applied Life Science (BK21), PMBBRC, Gyeo1

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2

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ngsang National University, Jinju 660-701, Korea; JGreen Inc., Department of Research & Development, Changnyeong 635-806, Korea 2

A novel endophytic bacterium (Nocardioides sp. YC6903) was isolated from a halophyte (Carex heterolepis L.). Radish seedlings were inoculated with YC6903 and grown gnotobiotically on MS media for 14 days in a growth chamber. After 7 days incubation, inoculated plants were more resistant to salt stress than control plants. In pot test, this strain also showed growth promotion of radish seedlings and salt resistance significantly in the presence of 150 mM NaCl for 4 weeks. The 16s rRNA gene sequence of the strain YC6903 had 97% similarity with the N. pyridinolyticus OS4 . DNA-DNA relatedness between T

causal agent of soft-rot disease of many agriculturally important crops. Here, we analyzed the role of the rpoE which encodes an alternative sigma factor (σ ) on virulence and stress response of D. dadantii 3937. Phenotypic analysis revealed that the mutant produce a reduced level of pectate lyase compared to that of the parental strain. Virulence of the mutant was reduced in Chinese cabbage compared to that of the parental strain. The mutant showed increased susceptibility to antimicrobial peptide salmon protamine and oxidative stress. These results suggest that the translational product of D. dadantii 3937 rpoE gene plays important role in its virulence and stress response. E

B-16

The pathogenicity island of Erwinia pyrifoliae WT3, Japanese Erwinia EjP617 and its relatedness to Erwinia amylovora pathogenicity island (PAI). Shree Prasad Thapa , Eun Chang Kim , Duck 1

2

Hwan Park , Sae Youll Cho , Jang Hyun Hur and Chun Keun Lim . 2

2

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Division of Biological Environment, College of Agriculture and Life Sciences, Kangwon National University, Chuncheon 200-701, Korea; Division of Bio-resources Technology, College of Agriculture and Life Sciences, Kangwon National University, Chuncheon 200-701, Korea 1

2

The Japanese Erwinia spp. first isolated in pears from Hokkaido, Japan, was tentatively identified as E. amylovora and later shown to be related to E. pyrifoliae. To describe the relatedness between E. pyrifoliae and Japanese Erwinia Ejp, we have sequenced PAI of E. pyrifoliae WT3 and Japanese Erwinia Ejp617 and then compared them with E. amylovora PAI. The 114kb gene segment of E. pyrifoliae has all the characteristic features of PAI and has been designated „Pyrifolian pathogenicity island. The pyrifolian PAI carries 38kb hrp gene cluster comprising eight transcriptional units and nine hrc genes along with 78 open reading frames (ORFs). The gene segments of Japanese Erwinia Ejp617 are also similar in size and collinear to those of the pyrifolian PAI. Genomic comparison of E. pyrifoliae WT3 to Japanese Erwinia Ejp617 and E. amylovora revealed that both WT3 and Ejp617 are highly homologous (>97% homology) to each other. Although the hrp/hrc cluster of E. amylovora was highly homologous to those of WT3 and Ejp617, a striking difference was found in the regions upstream of rlsA containing 55 ORFs (IT region of E. amylovora) and downstream of hrpA (orf23-orf29) operon. These results, thus, indicate that the Japanese Erwinia Ejp617 is similar to E. pyrifoliae strain rather then E. amylovora.

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B-17

Characterization of Erwinia rhapontici isolated from Asian pear by 16S rRNA and 16S-23S intergenic spacer regions and sequencing of hrpN gene. Shree Prasad Thapa , Eun Chang Kim , 1

2

Duck Hwan Park , Saeyoull Cho , Jang Hyun Hur and Chun Keun Lim . Division of Biological Environment, College of Agriculture 2

2

2

1

1

and Life Sciences, Kangwon National University, Chuncheon 200701, Korea; Division of Bio-resources Technology, College of Agriculture and Life Sciences, Kangwon National University, Chuncheon 200-701, Korea

B-19

Mutations in ColS/ColR Reduce Virulence and Oxidative Stress Resistance of Xanthomonas oryzae pv. oryzae KACC10859. Jung-

Hee Cho , Min-Jung Jeong , Woo-Jae Kim and Jae-Soon Cha . 1

1

2

1

Department of Plant Medicine, Chungbuk National University, Cheongju 361-763, Korea; Honam Agricultural Research Insstitute, RDA, Iksan 570-080, Korea 1

2

2

Four strains were isolated from pear cankers of Asian pear trees Pyrus pyrifolia, on the vicinity of Chunchon, Korea and were characterized on the basis of physio-chemical and molecular analyses using 16S rRNA gene, 16S-23S ITS region and hrpN gene. All four strains were similar to the representative strain tested physio-chemical experiments, however, none of the strains in the present study produced pink pigment a typical characteristic of E. rhapontici. Phylogenetic analysis using 16S rRNA and ITS region sequences data also revealed that they formed a discrete clade with high bootstrap value close to E. rhapontici and integrated it to none-soft rot group (or amylovora group) different from the carotovora group previously reported by Dye (Dye 1969a). The hrpN of 2OP2 strain, which is representative of four isolated strains, revealed a high homology (99.3%) with the hrpN of E. amylovora, whereas it differed in three single nucleotide polymorphisms (SNPs) substitution at 421bp, 491bp and 980bp, and an indel at position 1117bp. The molecular mass of HrpN protein was approximately 40kDa and lacked two amino acid fragments at the C-terminal region, which differentiated it from HrpN protein. Artificial inoculations on immature pear fruits and shoot of Pyrus pyrifolia showed pathogenicity, thus evidencing it as one of the prospects for causing disease in pear. Er

Ea

B-18

Occurrence of onion bacterial rot disease caused by Bacillus velezensis. Sun Kyoung Hwang¹, Chang Ki Beak , Hee Dae Kim , 2

1

Hee Young Jung². ¹Onion Research Institute, Gyeongnam Agricul-

tural Research and Extension Services, Changnyeong, 635-821, Korea; ²College of Agricultural and Life Sciences, Kyungpook National University, Daegu 702-701, Korea

In 2008, a bacterial rot of onion (Allium cepa L.) bulbs was observed in several low-temperature warehouses in Changnyeong-gun. The bacteria were isolated from lesions with conventional plating methods on NA (Nutrient Agar) medium and we obtained 8 bacterial isolates from rotten onion bulbs collected in several independent places. All isolates showed the pathogenicity on excised piece of onion scale and abundant bacterial growth was observed on the inoculated onion pieces. The optimum temperature for the growth and colony formation was 30 C on NA medium. The bacterial spores were single rodshape, 2.7~3.9×0.58~0.65. The sequence analysis of 16S rDNAs showed that all isolates were perfectly equal to each other and a few of the Bacillus species. The gyrA gene sequences known as the fast differentiation method of B. species were very similar(>99.9%) to B. velezensis. On the basis of these data we show that the isolated bacterium is identified as B. velezensis. This is the first report of bacterial rot of onion caused by B. velezensis in Korea.

Mutations in rpfC and rpfG homologs in Xanthomonas oryzae pv. oryzae (Xoo) reduced virulence in our previous study. Microarray analysis of the rpfC− mutant was carried out to screen the RpfC regulons in Xoo. One of the significantly down regulated genes in the rpfC− mutant was colS/colR genes that were known as a two-compo-

nent signal transduction system involved in competence of root-colonizing in Pseudomonas fluorescens. Promoter activity of colS/colR was reduced in the rpfC− mutant, which suggests that expression of colS/colR is controlled by RfpC in Xoo KACC10859. Marker exchange mutations in colS/colR reduced the lesion length to about 65% of wild type's. Virulence determinants such as cellulase, motility, xylanase and EPS were significantly reduced in the mutants. Also, the mutations made the strains more sensitive to hydrogen peroxide and produce more siderophore. Complementation of the mutation restored the mutants phenotypes. These results suggest that colS/colR are involved in virulence expression and resistance against oxidative stress in Xanthomonas oryzae pv. oryzae KACC10859.

B-20

Effect of recombinant N-acyl Homoserine Lactone Lactonase gene on Erwinia infection. Kim Cuc Nguyen Thi , Mi-Sun Yoo , Il1

1

Wook Kim , Choong-Hyo Yun * and Byoung-Su Yoon *. 1

2

1

Department of Life Science, Graduate School of Kyonggi University, Suwon, 443-760, Korea; National Academy of Agricultural Science, Suwon, 441-707, Korea 1

2

N-acyl Homoserine Lactone Lactonase (AHLase) produced by Bacillus thuringiensis was reported to be able to inhibit Quorum-sensing. In this study, we carried out cloning and expression of AiiA gene encoding for AHLase and expressed recombinant protein showing suppressive activity on Ewrinia virulence. AiiA gene originated from KACC12082 was amplified, cloned and sequenced. It showed 99% and 100% nucleotide and amino acid sequence identification with AF478052 (Bacillus thuringiensis serovar kyusuensis), respectively. The gene was inserted into pET32a(+) vector to express the protein. This protein was extracted and used for bioassay. In the assay, recombinant AHLase protein showed high protection on potato slice and Chinese cabbage from the attack of plant pathogenic Erwinia caratovora. The recombinant protein was successfully expressed and had the protective ability against Erwinia virulence. For the future plan we will develop a efficient system to purify pure recombinant AHLase.

o

B-21

Specific detection of Xanthomonas oryzae pv. oryzae by SYBR Green Real-Time PCR using pathovar specific primers based on rhs family gene sequences. Min Seok Cho, Man Jung Kang, Hong Il

Ahn, Jang Ho Hahn, Hyung-Jin Baek, Young Wang Na, Yu Mi Choi, Tae San Kim and Dong Suk Park*. National Academy of Agricultural Science, Rural Development Administration, 441-707 Suwon, Korea

Abstract of Presentations The present study describes PCR assays to detect bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae in rice. Two sets of PCR primer were developed to amplify gene required for a rhs family gene homologous to RhsA, cell envelope biogenesis, outer membrane. Only each PCR product of 114 bp and 297 bp was respectively amplified with the two primer sets, XOOP1F/XOOP1R and XOOP3F/ XOOP3R, from 9 isolates of X. o. pv. oryzae which were collected from several countries. In real time PCR, this primer pair was able to detect at least 5 pg/ul equivalents of genomic DNA. Moreover, the primer pair was able to detect the pathogen in artificially inoculated leaf of rice. Besides, dot blot analysis showed the same result with probe of the PCR product. In this study, SYBR green real-time and conventional PCR assay method can be used for the rapid detection of X. o. pv. oryzae, and will potentially simplify and facilitate the diagnosis and monitoring of this pathogen.

B-22

272

native sigma factor (σ ) on virulence and stress response of D. dadantii 3937. Phenotypic analysis revealed that the mutant produce a reduced level of pectate lyase compared to that of the parental strain. Virulence of the mutant was reduced in Chinese cabbage compared to that of the parental strain. The mutant showed increased susceptibility to antimicrobial peptide salmon protamine and oxidative stress. These results suggest that the translational product of D. dadantii 3937 rpoE gene plays important role in its virulence and stress response. E

B-24

Occurrence of crown gall on Korean black raspberry caused by Agrobacterium tumefaciens. Young Kee Lee , Inn Shik Myung , Jae 1

1

Hyeon Park , Chang Seok Kim , Sung Kee Hong , Hyo Won Choi , Sang Yeob Lee and Wan Gyu Kim . Agricultural Microbiology 1

2

1

1

1

1

1

Division, National Academy of Agricultural Science (NAAS), Rural Development Administration (RDA), Suwon 441-707, Korea; Organic Agriculture Division, NAAS, RDA, Suwon 441-707, Korea

2

Pathogenicity and molecular characterization of endo-glucanase gene cluster in Xanthomonas oryzae pv. oryzae. Jae-Won Kim , 1

Ulambayar Temuujin , Jong-Kun Kim , Jeong-Gu Kim , ByoungMoo Lee and Hee-Wan Kang . The Graduate School of Biotechnol1

1

2

1

2

1

ogy and Information Technology, Hankyong National University, Anseong 456-749, Korea; National Academy of Agricultural Science, Suwon 441-707, Korea 2

The xoegl1, xoegl2 and xoegl3, endo-glucanase genes were present in Xanthomonas oryzae pv. oryzae genome as a cluster. Each egl gene was PCR amplified and cloned in T-vector and transposons were inserted in the genes. Transposon inserted xoegl genes were marker exchanged with those of wild type strain (KACC 10859). The their marker exchanged events were confirmed by PCR and Southern hybridization. The mutants, xoegl1::Tn and xoegl2::Tn were nonpathogenic on rice variety, Milyang23, but xoegl3::Tn was pathogenic on the rice like wild type strain. Peptide consisting of 20 amino acids was synthesized from N-terminal of Xoegl1 protein and antibody was raised against the peptide. xpsF (type II secretion system) mutant strain (Xom31) and hrp X mutant (hrpX::Tn5), xoegl1::Tn and wild type strain were cultured in hrp inducing XOM2 medium and their extracellular protein patterns were compared on SDS-PAGE by silver staining. Western blotting analysis using anti-Xoegl1 peptide were further done on the SDS-PAGE. xoegl1::Tn and xoegl2::Tn exhibited identical cellulase activity to wilde type strain KACC10859 forming halo zones on carboxymethylcellulose (CMC) medium, while xpsF::Tn and hrpX::Tn mutant strains did exhibit reduced cellulase activity on the medium.

Crown gall symptoms on roots and stems of Korean black raspberry, Rubus coreanus Miq., were observed in Sunchang, Korea in 2006 and in Jeongup in 2008. Tumors on the plants were semi-round with rough surface texture of whitish or dark brown color. A total of 8 isolates obtained from the gall tissues were identified as Agrobacterium tumefaciens based on cultural, biochemical and physiological characteristics. Their colonies were convex, glistening, circular with an entire edge, and white to tannish-cream in color on PDA plus 0.5% CaCO . The bacteria were gram-negative, aerobic growth, coloniesmucous on YDC, urease-positive, oxidase-positive, growth on D1M agar, motility at pH 7.0, growth in 2% NaCl, growth at 35 C, and alkaline on litmus milk. The strains produced 3-ketolactose, acid from ethanol, melezitose, sucrose and propionic acid, and a reddish brown pellicle at the surface of the ferric ammonium citrate broth. Young Korean black raspberry plants inoculated with the bacteria developed typical galls within two to four weeks. Seedlings of tomato also produced typical galls two to four weeks after inoculation. This is the first report that Agrobacterium tumefaciens causes crown gall of Korean black raspberry in Korea. 3

o

B-25

Crown gall of peach caused by tumorigenic Agrobacterium rhizo-

genes. Young Kee Lee , Jae Hyeon Park , Inn Shik Myung , Jae Soon Cha , Sang Yeob Lee , Sung Kee Hong , Hyo Won Choi and Wan Gyu Kim . Agricultural Microbiology Division, National Academy 1

2

1

1

1

1

1

1

1

of Agricultural Science, Rural Development Administration, Suwon 441-707, Korea; Department of Plant Medicine, Chungbuk National University, Cheongju 361-763, Korea 2

B-23

Analysis of the role of rpoE gene on virulence and stress response of Dickeya dadantii 3937. Md. Shahinur Kabir , Hisae Hirata and 1

2

Shinji Tsuyumu . Institute for Genetic Research and Biotechnology, 1,2

1

836 Ohya, Suruga-ku, Shizuoka 422-8529, Shizuoka University, Japan; Faculty of Agriculture, 836 Ohya, Suruga-ku, Shizuoka 4228529, Shizuoka University, Japan 2

Dickeya dadantii 3937 (formerly Erwinia chrysanthemi 3937) is the

causal agent of soft-rot disease of many agriculturally important crops. Here, we analyzed the role of the rpoE which encodes an alter-

Occurrence of crown gall on roots of peach, Prunus persica var. vulgaris Maxim., was observed in Suwon, Korea in 2000 and in Iksan in

2001. Pathogenicity of 11 isolates obtained from the gall tissues was confirmed on stems of tomato by artificial inoculation. All the bacteria were identified as Agrobacterium rhizogenes and A. rhizogeneslike bacteria based on cultural, biochemical and physiological characteristics, and 16S rDNA sequences. Their colonies were convex, glistening, circular with an entire edge, and white to tannish-cream in color on PDA plus 0.5% CaCO . The strains classified as 4 different types based on biochemical and physiological characteristics. All the 3

273


strains were positive for aerobic growth, urease, growth on D1M agar, acid production from sucrose, oxidase reaction, and citrate utilization, but different for other tests. 16S rDNA gene sequence analysis revealed that the 11 isolates belonging to A. rhizogenes and A. rhizogenes-like bacteria. This is the first report that Agrobacterium rhizogenes causes crown gall of peach in Korea.

B-26

Crown gall of oriental melon caused by Agrobacterium larrymoorei. Young Kee Lee , Dong Geun Kim , Inn Shik Myung , Jae 1

2

1

Hyeon Park , Sung Kee Hong , Sang Yeob Lee , Wan Gyu Kim and Jae Soon Cha . Agricultural Microbiology Division, National Acad1

1

3

1

1

1

emy of Agricultural Science, Rural Development Administration, Suwon 441-707, Korea; New Material Research Center, Uiseng 760803; Department of Plant Medicine, Chungbuk National University, Cheongju 361-763, Korea 2

3

Occurrence of crown gall on lower stems of oriental melon, Cucumis melo var. makuwa Makino, was observed in Seongju, Korea in 2000 and 2001. Tumors on the plants were semi-round with rough surface texture of whitish or brown color. Four isolates obtained from the gall tissues were identified as Agrobacterium larrymoorei based on cultural, biochemical and physiological characteristics, and 16S rDNA sequences. Their colonies were convex, glistening, circular with an entire edge, and white to tannish-cream in color on PDA plus 0.5% CaCO . The bacteria were gram-negative, aerobic growth, coloniesmucous on YDC, urease-positive, oxidase-positive, growth on D1M agar, motility at pH 7.0, growth in 2% NaCl, and acid production from sucrose. The strains classified as 3 different types based on biochemical and physiological characteristics. 16S rDNA gene sequence analysis revealed that the 4 isolates belonging to A. larrymoorei. Pathogenicity of the bacteria obtained from the gall tissues was confirmed on stems of oriental melon and tomato by artificial inoculation. This is the first report that Agrobacterium larrymoorei causes crown gall of oriental melon in Korea. 3

Myoviridae, Phodoviridae, and Sipoviridae, and all three families are placed in the order Caudovirales. Since Sipoviridae were found in all six regions where bacteriopages were isolated, DNA from different Sipoviridae had been characterized.

B-28

Identification and Cloning of the Receptor gene for Target Recognition by Carocin D from Pectobacterium carotovorum subsp. carotovorum Pcc3. Eunjung Roh, Tae-Ho Park, Dong Hwan Lee,

Sannyu Jee, Dooho Kim and Sunggi Heu. Microbial Safety Division, National Academy of Agricultural Science, RDA 441-707, Suwon, Korea

The carocin D (caroDK) is a novel bacteriocin produced by P. carotovorum subsp. carotovorum Pcc21. A mutant strain of Pectobacterium carotovorum subsp. carotovorum Pcc3-M1, selected on the basis of its resistance to carocin D, reveals distinct functional alternations in unspecific OmpN porin. OmpN genes isolated from not only sensitive wild type P. carotovorum subsp. carotovorum Pcc3 strains but also resistant other Pectobacterium strains had been cloned and characterized. The sensitivity against carocin D of P. carotovorum subsp. carotovorum Pcc3-M1 was recovered with the wild-type outer membrane gene, pLOMP3. However, P. carotovorum subsp. carotovorum Pcc3M1 transformed with ompN genes isolated from resistance strains were resistanct as before. Alignment of the OmpN protein sequence of PCC3 strain with selected sequence from sensitive and resistance Pectobacterium strains was revealed significant variations on the cell surface exposed regions.

C-1

Biological control of grapevine crown gall by nonpathogenic Rhizobium (=Agrobacterium) vitis strain VAR03-1. Akira Kawaguchi , 1

Koji Inoue , Yuki Ichinose and Hideo Nasu . Agricultural Experi1

2

3

1

ment Station, Okayama Prefectural General Agriculture Center, 1174-1, Kouda-oki, Akaiwa-city, Okayama, 709-0801, Japan; The Graduate School of Natural Science and Technology, Okayama University, 1-1-1 Tsushima-naka, Okayama 700-8530, Japan; Zen-noh Okayama, 9-18-201, Togiya-cho, Okayama, 700-0826, Japan 2

B-27

Distribution and characterization of bacteriophages of Pectobacterium caratovorum subsp. carotovorum isolated from six different regions of Korea. Sannyu Jee, Eunjung Roh, Tae-Ho Park, Dong Hwan Lee, Kyusuk Jung, Dooho Kim and Sunggi Heu. Microbial

Safety Division, National Academy of Agricultural Science, RDA, Suwon 441-707, Korea

Soft rot is one the most important disease that causes high economic loss in Korean vegetable cultivation. As a pathogen of soft rot, Pectobacterium carotovorum subsp. carotovorum is able to infect growing plants as well as the harvested crops. Since the use of phages for disease control is a fast expanding potential to replace the chemical control, 124 bacteriophages were isolated from the soil samples collected in Pyeongchang, Taebaek, Gochang, Muan, Haenam and Jeju, where are the places for the most Chinese cabbage cultivation area. Using selected 50 different P. carotovorum subsp. carotovorum strains isolated in Korea, the host ranges of phages were tested. For the purification of bacteriophages, bacteriophage lysates were purified with density gradient centrifugation using cesium chloride. Based on morphotypes, purified bacteriophges were classified under three families,

3

Crown gall of grapevine, caused by Rhizobium (=Agrobacterium) vitis (Ti), is the most important bacterial disease of grapevine throughout the world. The screening tests of biological control agent in nonpathogenic R. vitis were carried out, and nonpathogenic R. vitis strain VAR03-1 was selected as biological control agent against grapevine crown gall. In a test with a 1:1 cell ratio of pathogen/nonpathogen on stems of tomato, sunflower, and grapevine, strain VAR03-1 was effective in reducing the incidence of tumor formation on grapevine and reduced tumor size. Strain VAR03-1 was bacteriocinogenic, causing an inhibition zone against tumorigenic R. vitis strains on YMA medium. In biological control tests, using the method of planting grapevine seedlings in the soil infested with some tumorigenic strains, treatment of grapevine roots with strain VAR03-1 (adjusted to 10 cells/ml and soaked for 1 h) reduced percentage of plants with tumors and controlled crown gall of grapevine seedlings. In a field test, furthermore, soaking-treatment of grapevine roots with strain VAR03-1 reduced percentage of plants with tumors and controlled crown gall of grapevine seedlings. VAR03-1 established popu9

Abstract of Presentations lations averaging 10 CFU/g root in the rhizosphere of grapevine and persisted on roots for two years. When roots of tomato and rose were soaked in a cell suspension of strain VAR03-1 before planting in soil infested with R. radiobacter (Ti) and R. rhizogenes (Ti), respectively, treatment with VAR03-1 reduced the number of plants with tumors. VAR03-1 was bacteriocinogenic, producing a halo of inhibition against those three species of tumorigenic Rhizobium. Thus, the sufficient results that VAR03-1 was an effective bacterial agent to control grapevine crown gall using this method were obtained. 6

C-2

Ecologically Sound Approaches for the Management of Anthracnose Disease of Noni. Manjunath Hubballi *, T. Anand , T. 1

1

Raguchander , S. Nakkeeran and R. Samiyappan . Department of 1

2

1

1

Plant Pathology Oil Seed Research Station, Tamil Nadu Agricultural University, Coimbatore-641003, Tamil Nadu, India 2

Noni (Morinda citrifolia) is a small, fruit-bearing, evergreen shrub or tree and is grown throughout the tropical countries for medicinal purpose. Anthracnose disease caused by Colletotrichum gloeosporioides (Penz) Sacc. is a major limiting factor in all Noni growing areas. Various isolates of bioagents were screened against the anthracnose pathogen. Pseudomonas fluorescens isolates TDK1, Bacillus subtilis isolate SVPR4 and Trichoderma viride isolate TV1 were found to be highly effective in reducing the mycelial growth of C. gloeosporioides under in vitro. Hence, these effective isolates were used for the development of vermicompost and neem cake based and liquid bioformulations. The bioformulations were tested individually and in combination against the incidence of anthracnose disease both under glass house and field conditions. Soil application of vermicompost and neem cake based bioformulation mixture (TDK1+SVPR4+TV1) and foliar application of liquid formulation mixture (TDK1+SVPR4+TV1) were highly effective in reducing the disease incidence. Further the combination treatments increased the activities of various defense enzymes viz., peroxidase, polyphenol oxidase, Catalase, phenylalanine ammonia lyase and phenolics in noni plants against the pathogen. The mechanism associated with reduced incidence of anthracnose may be induction of defense mechanism in plants.

C-3

Screening of potential antagonistic bacteria for the biological control of different fungi. Kabir Lamsa , Sang-Woo Kim, Jin-Hee Jung l

and Youn-Su Lee. Division of Bio-Resources Technology, College of Agriculture and Life Sciences, Kangwon National University, Chuncheon 200-701, Korea

The study was conducted to select potential rhizobacteria for the control of three different fungi i.e. Fusarium oxysporum, Sclerotinia homeocarpa and Phytopthora infestans. These antagonistic bacteria were selected through the in vitro dual culture method. Most of the antagonists showed varying levels of antagonism in the dual culture assay. Through the sequential screening of different isolates of bacteria, eleven different kinds of bacteria were found to be effective towards the control of these fungi in vitro. Among them, seven bacteria showed antagonism towards S. homeocarpa and six bacteria showed inhibition towards P. infestans, whereas, only two bacterial isolates showed antagonism for the control of F. oxysporum. Among

274

the bacterial strains, AB-9 showed the highest inhibition rate (i.e. 45.30%) towards S. homeocarpa and with 48.83% of inhibition rate, AB-9 showed efficiency for the growth control of F. oxysporum. But in case of two different isolates, 1306 and KACC 40178 of Phytopthora infestans, two different bacterial strains, AB-15 and AB-14 respectively showed 58.13% of inhibition. Further identification of selected bacteria and the field experiments are still on the way.

C-4

Isolation and characterization of plant growth promoting bacteria from fermented bovine products showing plant growth promotion and antifungal activity. Rangasamy Anandham *, Yong Ki 1

Kim , Hyeong Jin Jee , Sung Jun Hong , Hang Yeon Weon and Soon Wo Kwon . Organic Agriculture Division, Korean Agricultural Cul1

1

2

1

2

1

2

ture Collection, National Academy of Agricultural Science, Rural Development Administration (RDA), Suwon 441-707, Korea

Plant diseases reduce yield and productivity of several economical crops all over the world. Resistant plant cultivars, cultural and chemical application are routinely used to provide disease control. Available chemical fungicides are often expensive and also have adverse effect on human beings. Therefore, biological control appears to constitute an alternative strategy for controlling diseases. In search of effective bio-control and plant growth promoting bacteria, totally 97 bacteria were isolated from fermented bovine products (dung, milk, urine and yogurt) and characterized through 16S rRNA gene sequence analysis. Of which, initially functional characterization was performed only for 14 bacteria. Bacillus subtilis subsp. subtilis M1T2B8c, Bacillus amyloliquefaciens M1T2B8z, Bacillus pycnus M2T8B11, Bacillus circulans M3T2B1, Bacillus atrophaeus M3T4B2, Bacillus stratosphericus M3T8B15, Paenibacillus stellifer M3T4B6, Paenibacillus castaneae M2T3B4, Pseudomonas stutzeri M1T8B15, Pseudomonas xanthomarina, M1T8B19, Pseudomonas libanensis M2T10B1, Pseudomonas gessardii M2T10B2, Azospirillum lipoferum M2T2B2 possessed multiple plant growth promoting traits (N2 fixation, ACC-deaminase activity, tri-calcium phosphate solubilization, chitinase, pectinase, cellulase, indole acetic acid and siderophore production). Sulfur oxidation activity was only observed in Bacillus amyloliquefaciens M1T2B8z and Paenibacillus stellifer M3T4B6. All the tested strains produce volatile antibiotics (except Azospirillum lipoferum and Bacillus circulans) and control the Alternaria solani, Fusarium sp., Rhizoctonia solani, Bipolaris sp., Colletotrichum sp., Curvularia sp. Botrytis sp. However, in dual plate assay the tested bacteria offer resistant against only minimum numbers of aforesaid pathogens. Of which Bacillus atrophaeus M3T4B2 inhibited the growth of Alternaria solani (1.3 cm), Rhizoctonia solani (0.2 cm), Bipolaris sp. (1.3 cm), Colletotrichum sp. (1.7 cm), Curvularia sp. (0.4 cm), Fusarium (0.9 cm) and Botrytis sp. (1.3 cm). Further, their potential to promote plant growth and resistant to powdery mildew were evaluated in green house experiments with tomato. The results revealed that all the tested bacteria promote plant growth and significantly improved dry mass accumulation of plants.

C-5

The strategy of Blumeria graminis f. sp. hordei for surviving after failure of first infection. Koreyuki Sugai , Hiromi Masaoka , Karma 1

1

Penjore , Yupa Hanboonsong , Masamichi Nishiguchi and Naoto 2

3

1

275


Yamaoka *. Laboratory of Plant Pathology, Faculty of Agriculture, 1

1

Ehime University, Matsuyama 790-8566, Japan; Faculty of Agriculture, College of Natural Resources, Lobesa, Bhutan; Laboratory of Entomology, Faculty of Agriculture, Khon Kaen University, Khon Kaen 40002, Thailand 2

3

ellum SKT-1(Ecohope), a microbial pesticide. Kenichi Kida , Kazuo Kumakura , Satoshi Watanabe , Takeshi Ichikawa and Makiichi Takagaki . Life Science Research Institute, Kumiai Chemical 1

2

1

2

3

1

Industry Co., Ltd., Kikugawa, Shizuoka, 439-0031, Japan; Kumiai Chemical Industry Co., Ltd., Taito-ku, Tokyo 110-8782, Japan; Shizuoka Agricultural Experimental Station, Iwata, Shizuoka 4260905, Japan 2

3

The powdery mildew fungus of barley, Blumeria graminis f. sp. hordei (Bgh), is an obligate parasite. If the fungus fails to form a haustorium in a barley cell, it cannot absorb nutrients from the host cell and will die. To survive after failure of the first penetration, the fungus attempts a secondary attack from a second lobe of the appressorium. If the secondary attack is successful, the fungus can form hausutoria in barley cells and take nutrients, and then finally propagates, forming new conidia. However, a highly resistant condition is induced in the same host cell as a result of the failure of the first attack by Bgh. This condition makes it difficult for the fungus to attempt to penetrate the same cell that the first lobe fails to penetrate. However, when the fungus fails to penetrate with the first lobe, there is a high tendency for a second lobe to emerge on the side opposite the first lobe. This phenomenon means that a second lobe would penetrate a cell adjacent to the cell of the first attack when the appressorial germ tube (AGT) lies parallel to the border between host cells. Moreover, most of the AGTs reach the cell border even if the germlings exist in the center of the host cells and attempt to penetrate adjacent cells on which germlings exist. Hence, the result shows that the second lobes consequently escape the highly inaccessible state in the same cell enhanced by the failure of a prior attack by the first lobe.

C-6

Root colonization of Pantoea annatis strain B1-9 that promotes plant growth and yield of pepper. Su-Nam Kim, Tran Thi Phuong

Chi and Chang-Seuk Park. Institute of Agriculture & Life Sciences, Gyeongsang National University, Jinju 660-701, Korea

In order to select plant growth promoting rhizobacteria, we isolated several bacteria from various plant rhizosphere regions. The diluted solution of each isolated bacteria was applied in rhizosphere of pepper seedling to check plant growth promotion. Among them, we concentrate to bacteria which equipped an ability to promote plant growth and readily to colonize in plant root. The B1-9 which was first isolated from the region in Myunggi, Gyeongnam, promoted growth of pepper plants. In the seedling experiment, pepper plants with an application of B1-9 increased the plant height (6%), the number of leaves (20%) and the plant fresh weight (19%) as compared to control plants. In addition, the yield of pepper with a treatment of B1-9 was three times higher than that of control plants in the field experiment. The 16S ribosomal RNA gene sequences of B1-9 revealed that B1-9 is highly homologous to the genus of Pantoea annatis. In addition, we analyzed abilities of Pantoea annatis strain B1-9 in the production of phytohormones and siderophore as well as phosphorus solubilization and sulfur oxidation. Furthermore, we observed colonization pattern and density of Pantoea annatis strain B1-9 tagged with green fluorescent protein (GFP) in pepper seedlings using a confocal laser scanning microscope (CLSM).

C-7

Properties and biological control activities of Trichoderma asper-

Kumiai Chemical Industry Co., Ltd. and Shizuoka Prefecture Agricultural Experiment Station found Trichoderma asperellum SKT-1 (Ecohope®) that was a valuable fungal strain for the control of various seed-born diseases of rice. By soaking treatments of seeds with conidial suspension, T. asperellum SKT-1 was found to be highly effective against Bakanae disease caused by Gibberella fujikuroi, Bacterial seedling blight caused by Burkholderia plantarii, Bacterial grain rot caused by Burkholderia gulmae, and Bacterial brown stripe caused by Acidoyorax avenae. The suppressive effects of T. asperellum SKT1 on these rice seedling diseases were nearly equivalent to those of chemical fungicides such as ipconazol-copper for Bakanae disease and oxolinic acid or copper hydroxide for bacterial rice seedling diseases. The activities against Bakanae disease were not observed in treatments using autoclaved dead-conidia or water-extracted conidia, although they were observed in treatments using living conidia or conidia washed with sterilized water. In addition, in situ interactions between T. asperellum SKT-1 and G. fujikuroi on rice seeds using GFP-tagged transformations under confocal scanning laser stereomicroscopy. G. fujikuroi disappeared from embyos of rice seeds after treatment with SKT-1. Then, in dual culture, hyphae of G. fujikuroi was penetrated by hyphae of T. asperellum SKT-1, and degradation of the cell walls of G. fujikuroi was observed under the scanning electron microscope (SEM). These results suggested that mycoparasitism was the mode of action of T. asperellum SKT-1. Finally, T. asperellum SKT-1 caused no injuries to rice seedlings by seed treatments. Further, no effects were observed on the growth of 28 crops of 12 families in soil treatments and in 17 crops of 10 families in foliar treatments. We successfully developed T. asperellum SKT-1 as a microbial pesticide and registered it as an agricultural pesticide under the trademark “Ecohope®” in 2003 in Japan.

C-8

Screening and efficacy of the bacterium antagonistic to Korean oak wilt fungus Raffaelea querci-mongolicae sp. nov. Yong-Nam

Kwon, Kyung-Hee Kim , Sang-Hyun Lee , Sang-Tae Seo , HyeYoung Oh , Byeongjin Cha . Division of Forest Insect Pests and Dis1

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eases, Korea Forest Research Institute, Seoul 130-712, Korea; Department of plant medicine, Chungbuk National University, Cheongju 361-763, Korea 2

The Korean oak wilt disease spread and damaged Korean oak trees since late 2004 and is discussed as a major disease across the country these days. The symbiotic ambrosia fungus Raffaelea querci-mongolicae sp. nov. causes mortality of the mainly Quercus mongolica among oak trees. The pathogen is transmitted by Platypus koryoensis, which makes a specialized structure called mycangia as a way to transfer the fungus. In this research, bacteria were isolated from mycangia and their antagonistic activity to R. querci-mongolicae was tested. The dispersing adults of P. koryoensis were collected from tree attacked by them in several localities. Besides R. querci-mongolicae,

Abstract of Presentations various microorganisms were isolated from the mycangia of P. koryoensis, and their antagonistic activity was tested against R. quercimongolicae. In vitro tests, 4 isolates of microorganisms which strongly inhibit the growth of R. querci-mongolicae were isolated from the areas of relatively low pathogen isolation rate including Gwangju, Paju, Gwacheon, Cheongwon and Wanju. These isolates were identified as Tsukamurella tyrosinosolvens by showing 99% similarity with it in 16S rDNA sequence analyses.

C-9

Isolation and antifungal activity of polyacetylenic acids from Prunella vulgaris var. lilacina against phytopathogenic fungi. Mi-

Young Yoon , Gyung Ja Choi, Kyoung Soo Jang , Yong Ho Choi , Byung Jin Cha and Jin-Cheol Kim . Chemical Biotechnology 1,2

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Research Center, Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea; College of Agricultural and Life Sciences, Chungbuk National University, Cheongju, Chungbuk 361-763, Korea 2

In a search for plant extracts with in vivo antifungal activity against various plant pathogenic fungi, we observed that the methanol extract of Prunella vulgaris var. lilacina exhibits potent in vivo antifungal activity against Magnaporthe grisea (Hebert) Barr (rice blast) on rice plants, Botrytis cinerea Pers ex Fr (tomato grey mould), Puccinia recondita Rob ex Desm (wheat leaf rust) on wheat plants and Colletotrichum coccodes (Wallr) S Hughes (red pepper anthracnose) on pepper plants. Further solvent fractionation showed strong fungicidal activities in the n-hexane fraction against the plant diseases. We isolated two antifungal compounds by silica gel column chromatography, Sephadex LH-20 column chromatography, C column chromatography and C preparative HPLC. Structural elucidation of polyacetylenic acids were conducted by means of EI-MS, H-NMR, C-NMR, HSQC, H- H COSY and HMBC data. The two compounds were identified as octodeca-9,11,13-triynoic acid and transoctadec-13-ene-9,11-diynoic acid. The antifungal activities of the two compounds were first reported in this study. 18

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C-10

Temporal and Spatial Distribution of gfp Tagged Bacillus licheniformis N1 on Strawberry Plants. Hyoung Ju Lee, Hyun Gi Kong, Ki Hyuck Choi, Byung Ju Moon and Seon-Woo Lee. Department of Applied Biology, Dong-A University, Busan 604-714, Korea

Monitoring bacterial survival and distribution in natural environment is necessary to use various bacteria as biological control agents for plant diseases. This study describes an enhanced gfp based tagging system for biocontrol bacteria and the usage of the tagging system for B. licheniformis N1. Overlap extention PCR method is used for fusion between promoterless gfp gene and P43 promoter isolated from Bacillus subtilis. The construct was introduced into B. licheniformis and gfp expression was observed when the tagged bacteria were sprayed on various tissues of strawberry plants. The plasmid based construct in B. licheniformis N1 was more stable than chromosomal integration. Using the plasmid containing gfp tagged B. licheniformis N1, spatial and temporal distribution of the B. licheniformis N1 strain were investigated on strawberry plants using dilution plate counting over time on various tissues and observation with confocal laser scanning microscope (CLSM) both in controlled growth cham-

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ber and in plastic house. Gfp tagged B. licheniformis N1 was stably survived up to 20 days on strawberry plants such as leaves, petioles and crowns in growth chamber. However, the bacterial population was unexpectedly decreased in 5 days after application on strawberry plants in plastic house. CLSM analysis revealed that gfp tagged B. licheniformis N1 was colonized leaves, petioles and crowns of strawberry plants well and maintained green fluorescens for 3 days, 5 days, 7 days on strawberry leaves, petioles, crowns in growth chamber, respectively. Therefore, this study can recommend an efficient treatment schedule of biocontrol bacteria to control plant disease based on the analysis of bacterial temporal and spatial distribution on plants.

C-11

Identification of a Chromosomal Locus and Involved in Antifungal Activity of Burkholderia pyrrocinia CH-67. Kwang Youll Lee, Seon-Woo Lee and Byung Ju Moon. Department of Applied Biology, Dong-A University, Busan 604-714, Korea

Burkholderia pyrrocinia CH-67, identified by using 16S rRNA sequencing and recA sequences analysis, was developed as a bio-control agent against Fulvia fulva. This strain exhibits antifungal activity

against several phytopathogenic fungi. By using plasposon mutagenesis and Fulvia fulva spore screening, we select two antifungal deficient mutants (M193 and M208). DNA sequence analysis of the flanking region indicated that the plasposon insertion in M193 was in the region which shows similarity to a modular enzyme complex of non-ribosomal peptide synthetase. In another mutant (M208), the insertion was found to be in a different region from that of Mutant M193. Single insertion of the plasposon into those mutants was verified by Southern hybridization. In order to select an media for producing non-ribosomal peptide in B. pyrrocinia CH-67, different media like Kings B, Lactose agar, NA, PDA and TSA was tested. PDA medium was finally selected as an optimum production for non-ribosomal peptide. In mutant M208, the antifungal activity was restored by complementation of the full length of a putative kinase-like gene introduced in trans. Moreover, in the presence of 10 mM ADP or 10 mM ATP and AMP, the antifungal activity was also restored in the mutant M208. Identification and characterization of the chemical structure of non-ribosomal peptide from B. pyrrocinia CH-67 is in progress.

C-12

Isolation and characterization of Bacillus sp. JBC16 to control postharvest spoilage by green and blue mold rot of citrus fruits. Hyosong Nam, Sang-Mi Yu, Seralathan Kamala Kannan, Bong-Sik Yun, Seung-Moon Park, Kui-Jae Lee and Yong Hoon Lee*. Division

of Biotechnology, Chonbuk National University, Deokjin-dong, Jeonju, Jeonbuk 561-756, Korea

Green molds of citrus, caused by Penicillium digitatum, respectively, are major spoilage factors during storage of satsuma mandarin which is the major citrus crop of Juju province of Korea. To control the fungal pathogens using antagonistic bacteria, we screened bacterial isolates to select potential biocontrol agent to green and blue mold of the fruit. The selected bacterial strain JBC16 was identified as Bacillus sp. based on the 16S rDNA sequence analysis. The strain, Bacillus sp. JBC16 strongly inhibited development of P. digitatum on wounded

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mandarins, which was artificially inoculated with spores of the pathogens. The incidence of green mold was between 80% and 90% on non-treated fruits and the treatment with JBC16 decreased incidence to 20-30% (control efficacy of 70% to 80%). The bacterial strain strongly inhibited the mycelial growth of P. digitatum, P. italicum, P. expansum, Colletotrichum acutatum, Fusarium oxysporum, Botrytis cinerea, Diaporthe citri, and Alternaria alternata The Bacillus sp. JBC16 didnt produce siderophore and volatile organic compounds. The results indicated that the antifungal compound secreted by the strain was the major factor for the growth inhibition of the pathogens and control efficacy.

C-13

Suppression of postharvest green and blue mold disease on citrus fruit by treatment of Bacillus sp. JBC2. Hyosong Nam, Sang-Mi Yu, Gun Woong Lee, Jong-Chan Chae, Byung-Taek Oh, Kui-Jae Lee and Yong Hoon Lee*. Division of Biotechnology, Chonbuk National University, Deokjin-dong, Jeonju, Jeonbuk 561-756, Korea

Enormous economic losses are occurred by the spoilage of fungal pathogens during storage and distribution of citrus fruits. To develop alternative control measures using effective microorganisms, rhizospheric bacteria were isolated and screened to select effective antagonists against green and blue mold, which are major diseases limiting the extension of storage. The finally selected antagonistic bacterium JBC2, which was identified as Bacillus sp. based on 16S rDNA sequence analysis, suppressed the incidence of green and blue mold on wounded citrus fruits with inoculation of spore suspension of Penicillium digitatum and P. italicum. The strain, Bacillus sp. JBC3 strongly inhibited development of P. digitatum and P. italicum on wounded mandarins, which were artificially inoculated with spores of the pathogens. The incidence of green mold was between 80% and 90% on non-treated fruits and the treatment with JBC2 decreased incidence to 20-30% (control efficacy of 70% to 80%). The treatment with JBC2 decreased incidence of blue mold to 20-30% (efficacy of 70% to 80%) compared with non-treated incidence between 90% and 100%. The strain, Bacillus sp. JBC2, strongly inhibited the mycelia growth of fungal pathogens, such as P. digitatum, Botrytis cinerea, Diapothe citri, P. italicum, P. expansum, Alternaria alternata, Colletotrichum acutatum and Fusarium oxysporum. Fantastically, the secondary metabolites of Bacillus sp. JBC2 also strongly inhibited the mycelial growth of P. digitatum, P. italicum, and P. expansum. The results indicated that Bacillus sp. JBC2 and/or antifungal compounds

would be promising bio-control agent and/or compound(s) to postharvest diseases of citrus fruits.

duced by P. chlororaphis O6 that could be involved in fungal suppression include phenazines and pyrrolnitrin. To determine the effective compounds of P. chlororaphis O6, we used GacS mutant, RpoS mutant, Phz mutant, and PrnA mutant with two different growth media, such as mung bean agar and potato dextrose agar against various phytopathogenic fungi. P. chlroraphis GacS mutant, RpoS mutant, and PrnA mutant lost antifungal activities on mung bean agar medium, but GacS mutant and Phz mutant lost antifungal activities on potato dextrose medium against various plant pathogenic fungi. Biochemical and transcriptional analyses of P. chlrorophis O6 indicated that P. chlororaphis O6 produced pyrrolnitrin as a major antibiotic metabolites, while phenazine was produced mainly on potato dextrose medium. These results indicated that production of these antimicrobial agents was sensitive to the nutrients supplied to the bacterium and required functional GacS and RpoS regulation. These factors may in part account for the variability of biocontrol under field conditions.

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Enhanced Antiviral Activity to TMV by Fed-Batch Cultivation and Formulation Technique for Commercialization of Acinetobacter antiviralis sp. Nov. Nam Gyu Kim, Joung Woong Kim, Hyeong Min Kim, Cheol Jang and In Cheon Hwang*. Central Research Institute, Kyung Nong Co., Ltd., Gyeongju 780-110, Korea

Inhibitors of plant viruses have been studied from various plants, microorganisms, and some mushrooms but no commercial pesticide for plant virus control has been developed until now, because they has been limited for practical uses for control efficacy, mass production, stability in field condition and so on. The central research institute of Kung-Nong co., Ltd. has been developed pesticide for plant virus control. As a result, the culture filtrate powder from Acinetobacter sp. as KNF2022 showed very high antiviral activity to TMV, PepMoV, PMMoV and PVY infection. The type strain KNF2022 (=KCTC0699BPT) was identified as new species Acinetobacter antiviralis sp. Nov. This study was aimed to enhance the antiviral activity of KNF2022 through the fed-bath culture using organic acid as feeding source. Formulation technique of KNF2022 SM (World Star Eco ) was studied for cost reduction and commercialization. The antiviral activity in TMV local lesion host was evaluated by using the half-leaf method in tobacco plants (Nicotiana tabacum cv. Xanthinc). The control value of antiviral activity in KNF2022 SM (World Star Eco ) was increased 4.9 fold higher than batch culture technical at 500 dilution rate in half-leaf method. Increasing the antiviral activity of KNF2022 SM was due to increasing of cell density in fed batch culture and cell lysis in extraction by alcohol type solvents. Also the KNF2022 SM (World Star Eco ) has outstanding advantages such as cost reduction in comparison with KNF2022 WP (World Star). ®

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C-14

Metabolites from Pseudomonas chlororaphis O6 differentially inhibit growth of phytopathogenic fungi. D. H. Kim , S. H. Han , J. 1

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Y. Park , H. H. Im , A. J. Anderson , Y. C. Kim . Department of Plant Biotechnology, Chonnam National University, Gwangju, Korea; Department of Biology, Utah State University, Logan UT84322, UT, USA 1

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C-16

A volatile substance from Talaromyces sp. promotes plant growth and suppresses the development of diseases on several plants.

Yasuo Yamagiwa , Kazuhiro Toyoda , Yoshishige Inagaki , Yuki Ichinose , Mitsuro Hyakumachi and Tomonori Shiraishi . Laboratory 1

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Pseudomonas chlororaphis O6 suppresses growth of the wilt pathogen, Fusarium oxysporum f. sp. radicis-lycopersici, and the fungus causing wheat and barley scab, F. graminearum. Metabolites pro-

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of Plant Pathology and Genetic Engineering, Graduate School of Natural Science and Technology Pkayama University, Okayama, 7008530, Japan; Laboratory of Plant Pathology, Graduate School of 2

Abstract of Presentations Applied Biological Sciences, Gifu University, Gifu, 501-1193, Japan

A Talaromyces sp. promoting plant growth was isolated from an agricultural field at Okayama Pref. The strain FS2 was identified as T. wortomannii based on ITS1 sequence and morphology. FS2 enhanced seed germination, root elongation and leaf growth of Brassica. The leaf growth was also accelerated by volatile substance(s), that was excreted from FS2-culturing PDA. GC-MS analysis of the volatiles indicated that FS2 produced at least seven kinds of terpenoids. One of them was identified as b-caryophyllene that promotes the growth of leaf and stem of Brassica, Crucifer and Nicotiana. It also accelerates root development of Brassica Thus a part of the plant growth promoting effect of FS2 seems to be dependent upon this terpene. Interestingly, we also found that Brassicae, Crucifer and Nicotiana pretreated with this terpenne become tolerant to respective diseases caused by Colletotrichum higginsianum, C. lagenarium or Botrytis cinerea. Based on these findings, we discuss the availability of PGPF and its products for the court of crop cultivation.

C-17

Effects of Initial Inoculums of Paenibacillus polymyxa on Colony Formation and Starch-Hydrolytic Activity in Relation to Root Rot of Ginseng. Yong Ho Jeon , Sang Gyu Kim , Ingyu Hwang and 1,2

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Young Ho Kim . Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, Korea; KT&G Central Research Institute, Daejon 305-805, Korea 1

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Population changes, colony formation, starch hydrolytic- activity, ginseng root rotting of Paenibacillus polymyxa GBR-1 isolated from a rotten ginseng root were examined under in vitro and in vivo with the higher(1×10 colony forming units(CFU)/ml) and the lower (1×10 CFU/ml) inoculum levels. P. polymyxa GBR-1 showed a strong starch-hydrolytic activity on modified starch agar added with relatively low starch contents at given incubation temperatures (18 C and 23 C) only from the higher inoculum, producing the bacterial colonies about 9 times thicker than those formed from the lower inoculum. Light, scanning electron, and transmission electron microscopy showed that the thick colonies from the higher inoculum were filled with extracellular polymeric substances (EPS) in which a relatively small number of ovoid rod-shaped bacterial cells (mostly endosporebearing cells) were distributed. On the contrary, the thin colonies from the lower inoculum were composed of massive vegetative cells of a rectangular rod shape with minimum EPS. Fluorescent in situ hybridization (FISH) reveled that β-amylase gene was expressed only in the bacterial cells from the thick colonies formed from the higher inoculum, but not in those from the lower inoculum. The culture filtrate of the thick colonies produced a hydrolytic clear zone on modified starch agar, degraded starch granules in a various manner, and produced rot symptoms on ginseng root tissues. Biological properties of colony formation, starch hydrolysis, and ginseng tissue rotting of P. polymyxa GBR-1 are interrelated one another and influenced by the initial bacterial population but not by in situ and the final one. 8

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cultural Biotechnology, Seoul National University, Seoul 151-921, Korea

Bipolaris stem rot caused by Bipolaris cactivora is one of the serious diseases in cactus cultivation. In a pathogenicity experiment with the fungal conidia, we found that the conidial suspension sterilized at 121 C, 15 psi for 20 min showed an inhibitory effect on the disease development. In a greenhouse experiment, application of the sterilized conidial suspension (SCS) 2 days prior to the pathogen inoculation significantly reduced the disease severity, having only small brownish spots on cactus stems of Gymnocladium mianovichii and Hylocereus trigonus up to 4 weeks after inoculation. On the contrary, the cactus stems with pathogen inoculation only showed a severe stem rot from 5 days after inoculation and the whole stems were rotten 4 weeks after inoculation. For 6 isolates of B. cactivora, this disease-inhibitory effect of SCS held true against the stem rot caused not only by the same fungal isolates but also by the other different isolates. In another pathogenicity experiment using stem discs of H. trigonus and Cereus peruvianus, SCS also reduced the development of the stem rot, in which prominent boundary layers were formed beneath the inoculation sites of C. peruvianus stems. SCS was not and minimally effective in reducing the fungal mycelia growth and spore germination, respectively. Therefore, these results suggest that inhibition of the disease development in cactus stems treated with SCS may be derived mostly from the enhanced plant defense rather than inhibited pathogen growth and development. o

C-19

Biological control of Fusarium wilt of sweet potato by non-pathogenic Fusarium oxysporum SP5. Kee Don Han and Deuk Ho Kim.

Research Institute of Sungbo Chemicals Ltd., 455 Dukeundong Dukyanggu Goyangsi Gyunggi province,411-741, Korea

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C-18

Control of Bipolaris stem rot of cactus caused by Bipolaris cactivora with its sterilized conidial suspension. Min-Ok Choi, Min-

Hee Han, Hye-Young Kim and Young Ho Kim. Department of Agri-

Fusarium oxysporum f.sp. batats is a serious plant pathogenic fungus

that result in yield and quality losses of sweet potato. Recent works have suggested that the non-pathogenic isolates of Fusarium oxysporum could be suppressive effect on the Fusarium wilt of sweet potato. Fungal antagonists isolated from soil samples of sweet potato field in Goyang, Korea were evaluated for their potential as biological control agents. To select candidates of antagonist, isolates were inoculated on the sprout cuttings in humid plastic box test. For 7days after inoculation at 28 C, the 2 isolates selected in in vitro test were shown no pathogenic symptom and healthy sprouts. When they were previously inoculated using dipping the sweet potato spout cuttings into spore suspension (10 spore/ml) before being planted in the artificially infested soil, the SP5 was shown higher control value on the Fusarium wilt and fresh-weight than others in pot test at 30 days after planting. The SP5 isolates was identified as F. oxysporum based on morphological characteristics and 18S rRNA sequence analysis. These results indicated that F. oxysporum SP5 is considered to be applied for the practical biocontrol of Fusarium wilt of sweet potato using the pre inoculation on the sprout cuttings. o

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C-20

Hyperparasitism of Alternaria brassicicola by the biocontrol strain Streptomyces sp. MBCN152-1. Masafumi Shimizu , Miki1

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yasu Nakayama and Tomoko Suzuki . Laboratory of Crop Production and Ecology, Graduate School of Bioresources, Mie University, 1577 Kurimamachiya, Tsu 514-8507, Japan; Laboratory of Material Chemistry, Graduate School of Natural Science and Technology, Okayama University, 3-1-1 Tsushima-naka, Kita-ku, Okayama 7008530, Japan 1

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C-22

rDNA ITS sequence analysis and identification of Ampelomyces sp. isolate BSLAH16 hyperparasite of Powdery mildew. Ji Young

Kim , Wang Hyu Lee , In Young Choi , Ju Hee Kim and Sung Su Jung . Plant Medical Research Center, College of Agriculture and 1,2

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Life Science, Chonbuk National University, Jeonju 561-756, Korea; Chonbuk Agricultural Research and Extension Services, Iksan 570704, Korea 2

A non-antagonistic Streptomyces sp. strain MBCN152-1, isolated from surface-sterilized cabbage (Brassica oleracea L. var. capitata L.) collected from the experimental field of Mie University (Takanoo, Tsu city in Japan), has proved to be a potential biocontrol agent against Alternaria sooty spot, caused by Alternaria brassicicola, on cabbage seedlings. To elucidate the biocontrol mechanism of strain MBCN152-1, growth pattern of A. brassicicola inoculated onto cotyledons of cabbage seedlings pretreated with the strain was observed by light microscope. As a result, it was revealed that germ tube elongation and appressorial formation were significantly suppressed on the pretreated cotyledons. Next, spores of A. brassicicola were cocultured with strain MBCN152-1 on water agar and observed over time. Interestingly, mycelia of strain MBCN152-1 attached to the hyphae of A. brassicicola, coiling around them within a few days of inoculation. When those hyphae were stained with Calcoflour White M2R, bright fluorescent spots were observed in the regions of coiling. Appearance of intense fluorescence indicated the occurrence of localized cell wall lysis at sites of interaction between strain MBCN152-1 and the pathogen. Eventually, such parasitized hyphae were completely disintegrated about 1 week after incubation. Scanning electron microscopy revealed that hyphae and germ tubes of A. brassicicola on the surface of cotyledons were also heavily parasitized by strain MBCN152-1. In addition, high chitinase activity was detected when strain MBCN152-1 was co-cultured with A. brassicicola on ISP-9 medium supplemented with carboxylmethyl chitin, compared with low level found with the strain alone. These results suggests that hyperparasitism of strain MBCN152-1 may be associated with its biocontrol effect against A. brassicicola.

C-21

Characteristics of antagonistic bacteria to anthracnose of pepper, Colletotrichum acutatum. Won-il Kim, Sung-Jun Hong, Yong-Ki

Kim, Eun-Jung Choi and Hyeong-Jin Jee. Organic Farming Division, National Institute of Agricultural Science, Rural Development Administration, Suwon 441-707, Korea

Colletotrichum acutatum is major pathogen of pepper, causing eco-

nomically important losses in Korea. In order to control pepper anthracnose biologically, five antagonistic bacteria to Colletotrichum acutatum were selected as a biological control agent for pepper anthracnose. Among five selected antagonistic bacteria, Pseudomonas sp. U46-6 inhibited the mycelial growth of pathogens effectively. For more clear affiliation of the antagonistic bacteria, several biochemical tests were performed. In the biochemical tests, S11-10 showed the capability to produce IAA. Both U46-6 and U46-5 had ability to produce siderophore well. When the suspension of Pseudomonas sp. U46-6 was sprayed on pepper fruit after inoculation with C. acutatum, disease incidence of pepper anthracnose was decreased both wound and non-wound inoculation significantly. However, further studies are needed to test its effect of antagonism to pepper anthracnose in the field.

The results of these studies were partly presented in the autumn meeting of KSPP in 2008. Recently, BSLAH16 isolated Impatiens balsamina L. studied on rDNA ITS sequence analysis. Genomic DNA was extracted and rDNA ITS region was amplified by PCR with primer ITS1, ITS4. rDNA. ITS sequence of BSLAH16 was compared with other Ampelomyces sp. strains in GenBank by BLAST search in NCBI (National Center for Biotechnology Information). Size of entire rDNA ITS region which was amplified with primer ITS1, ITS4 for BSLAH16 was 570bp and yielded 449 aligned nucleotide positions for all species included in the alignment. The result compare with other 17 Ampelomyces sp. strains BSLAH16 shares 100% similarity with 4 spices which were clear identificion as a A. quisqualis isolate clear identification. Therefore BSLAH16 was clear identified as a A. quisqualis.

C-23

Establishment of effective treatment method of Oxyporus latemarginatus EF069 for the control of Chinese cabbage clubroot. Myoung Soo Park, Jiye Ahn, Gyung Ja Choi, Kyoung Soo Jang, Yong Ho Choi and Jin-Cheol Kim. Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea

Clubroot of Chinese cabbage, which is caused by Plasmodiophora brassicae, is the most serious disease in Korea. The objectives of this study were to investigate the effectiveness and effective treatment methods of Oxyporus latemarginatus EF069 for controlling Chinese cabbage clubroot. Wheat bran-rice hull cultures of EF069 into nonautoclaved soil as ratio of 1:19 was found to be more effective in reducing disease severity than other dosage in non-autoclaved soil and horticulture nursery medium. When non-autoclaved soil was mixed into horticulture nursery media at 0:10, 1:9, 2:8, 4:6, 5:5, 6:4, 8:2, 10:0, optimal blending ratios of non-autoclaved soil and horticulture nursery medium were 0:10 and 5:5 for suppression of clubroot disease and 8:2 for growth. O. latemarginatus EF069 effective controlled cabbage clubroot in non-autoclaved soil blended with 5:5 of autoclaved horticulture nursery media. EF069 showed higher control efficacy 86% for two- time applications of 5% (v/v) solid cultures of EF069 at sowing and transplanting into clubroot-contaminated soil. In the field, treatment of 5% (v/v) solid cultures of EF069 at sowing had 47.9% disease reduction compared with untreated control. These results indicated that O. latemarginatus EF069 could be used as a biological agent for cabbage clubroot.

C-24

Selection of basal liquid medium for the enhancement of disease control efficacy of Microbispora rosea subsp. rosea A073 against phytophthora blight. Jiye Ahn , Myung Soo Park , Gyung Ja 1,2

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Abstract of Presentations Choi , Kyoung Soo Jang , Yong Ho Choi , Jae Eul Choi and JinCheol Kim . Chemical Biotechnology Research Center, Korea 1

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Research Institute of Chemical Technology, Daejeon 305-600, Korea; College of Agricultural and Life Sciences, Chungnam National University, Daejeon 305-764, Korea 2

Microbispora rosea subsp. rosea A073 was selected as an effective

biological control agent of phytophthora blight. At first, the optimum agar medium for cellular proliferation of A073 was investigated. Among 6 agar media, corn meal agar medium was best for cellular proliferation, followed by oatmeal agar medium. After incubation on corn solid medium for 14 days at 30 C, the dried solid cultures, at 5%, were applied into horticulture nursery medium at different times. Application of solid culture 7 days before pathogen inoculation was most effective; the control value was 80%. The dried solid cultures showed dose-dependent response in the suppression of phytophthora blight. When the solid cultures were treated at 1%, 2%, and 5%, the disease severity in the plants treated at 5% was least, followed in order by 2% and 1%. However, the solid corn cultures sometimes hindered normal growth of red pepper plants. Therefore, we tried to determine the optimal basal medium for liquid culture. A073 was incubated in corn liquid medium consisting of 5 g of corn granules and 100 ml distilled water for 7 days at 30 C. After incubation, the fermentation broth was filtered through 8 folds of cheese cloth and then applied onto pots 7 days before inoculation. The fermentation broth of A073 also displayed dose-dependent response in reducing symptoms of phytophthora blight without any deleterious effects on red pepper plants. In order to determine the optimal liquid culture medium, we tested six cereals such as rice, wheat, barley, rye, oat and corn. Among the 6 fermentation broths, barley liquid culture suppressed most effectively the development of phytophthora blight, followed by rye and corn. The other three fermentation broths were not active. Further study on the determination of optimal composition of liquid medium containing barley as a basal substrate is in progress. o

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α-L-diginopyransyl-(1→4)-β-D-cymaropyranoside) were firstly reported in this study. Six compounds isolated from C. wilfordii roots were in vitro tested for their antimicrobial activities against six phyto-

pathogenic bacteria and six phytopathognic fungi. Cynauricuoside A and wilfoside C1G strongly suppressed bacterial growth of Ralstonia solanacearum, Magnaporthe grisea and Botrytis cinerea. Six pregnane glycosides were active to tomato gray mold except by wilfoside C1GG. Three compounds of caudatin glycosides effectively suppressed the development of barley powdery mildew, while kidjoranine glycoside series were virtually inactive. To determine optimal extraction solvent, C. wilfordii roots were extracted with various solvents and then the extracts were tested for their antifungal activities against seven plant diseases. As the results, the ethyl acetate extract showed the most potent in vivo antifungal activity. This study describes that extracts of C. wilfordii roots can be developed as natural fungicides.

C-26

Selection of commercial bio-fungicides for the control of anthrocnose on Panax ginseng. Yeoung-Seuk Bae, Byeong-Yeol Yeon, Ki-

Chun Park, Seung-Won Kang and Young-Chul Kim. Ginseng

Research Division, Department of Herbal Crop Research, National Institute of Horticultural & Herbal Science, Rural Development Administration, Bisanri 80, Eumseong, Chungbuk, 369-873, Korea

A total of 13 commercial bio-fungicides registered in Korea was screened against anthrocnose caused by Colletotrichum fuscum on Panax ginseng in shaded greenhouse conditions. In preliminary experiments, we selected 3 commercial bio-fungicide products, wettable powder (QST713WP) and suspension concentrate (QST713SC) of Bacillus subtilis isolate QST713 and wettable powder (DBB1501WP) of B. subtilis isolate DBB1501, showing promising control efficacy for further experiments. Optimal spray concentrations of selected products were 500X for QST713WP and QST713SC, and 1000X for DBB1501WP. DBB1501WP showed promising control effect at the inoculum concentration of 1×10 conidia/ml, while QST713WP and QST713SC showed the best control effect at the inoculum concentration of 1×10 conidia/ml. Furthermore, DBB1501WP effectively controlled the disease until 10 days after application. 6

Antifungal activity of six pregnane glycosides isolated from Cynanchum wilfordii roots against plant pathogenic fungi. Nam Hee Choi , Gyung Ja Choi , Kyoung Soo Jang , Yong Ho Choi , Myung Soo Park , Jae Eul Choi and Jin-Cheol Kim . Chemical Biotechnol1,2

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ogy Research Center, Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea; College of Agricultural and Life Sciences, Chungnam National University, Daejeon 305-764, Korea

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In the course of search for plant extracts with potent in vivo antifungal activity against 7 plant diseases caused by plant pathogenic fungi, we found that the extract of Cynanchum wilfordii roots effectively suppressed the development of various plant diseases. Six antifungal substances were isolated from methanol extracts of C. wilfordii roots by using bioassay-guided fractionation. Their chemical structures were determined to be six pregnane glycosides. Six pregnane glycosides were identified as wilfoside C1N, wilfoside K1N, cynauricuoside A, wilfoside C1G, wilfoside K1GG and wilfoside C1GG. Wilfoside K1GG (kidjoranine 3-O-β-D-glucopyranosyl-(1→4)-β-D-glucopyranosyl-(1→4)-α-L-cymaropyranosyl-β-D-cymaropyranosyl-(1→4)-α-Ldiginopyransyl-(1→4)-β-D-cymaropyranoside) and wilfoside C1GG(caudatin 3-O-β-D-glucopyranosyl-(1→4)-β-D-glucopyranosyl-(1→4)-α-L-cymaropyranosyl-β-D-cymaropyranosyl-(1→4)-

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Screening of potential biocontrol agents against Alternaria blight on Panax ginseng in Korea. Yeoung-Seuk Bae, Byeong-Yeol Yeon, Ki-Chun Park, Seung-Won Kang and Young-Chul Kim. Ginseng

Research Division, Department of Herbal Crop Research, National Institute of Horticultural & Herbal Science, Rural Development Administration, Bisanri 80, Eumseong, Chungbuk, 369-873, Korea

Alternaria blight is the most important disease in the cultivation of Panax ginseng causing stem blight from early May and leaf blight from late May in Korea. Total 56 Bacillus strains were isolated from ginseng roots and Korean traditional foods, such as soybean paste and Kimchi, and screened for potential biocontrol agents against Alternaria blight by anti-fungal test to various ginseng pathogens, and greenhouse bioassay. Eight isolates had strong anti-fungal activities against various ginseng pathogens. Among them, three Bacillus isolates

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showed the similar biocontrol efficacy than that of a registered commercial bio-fungicide, B. subtilis QST713 in laboratory or greenhouse test. Their possible mechanisms are discussed.

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Biological control of root-knot nematode using antagonistic bacterium Rahnella aquatilis No-16. Su Jin Lee, Yun Hee Seo, Ye-Lim

Jang and Young Ho Kim. Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, Korea

significantly influenced by infection pressure of powdery mildew on cucumber at the naturally infected greenhouse. Their therapeutic efficacies drastically decreased in proportion to the disease incidences of powdery mildew of cucumber, even though the biofungicides were consecutively sprayed two or three times.

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Biological Control of Fusarium Wilt to Lettuce by Bacillus subtilis GG95 in the Greenhouse. Hyun Ju Lee, Soon Sung Hong, Jin Young Kim, Jin Gu Lee and Sung Kee Kim. Gyeonggi-do Agricul-

Root-knot nematodes, Meloidogyne spp. are recognized as serious pests in cucumber production and it is very difficult to control the nematode disease using nematicides. Biological means is a good alternative for their control. In this study, 80 bacterial isolates obtained from soil and plant material sources were subjected to in vitro screening tests for the mortality of the root-knot nematode, M. incognita, out of which Rahnella aquatilis No-16 and Enterobacter spp. showed strong nematicidal activities. Especially, a combined treatment of R. aquatilis No-16 and stem bark of Cinnamomum loureirii (SBC) showed a greater nematode mortality of about 98% than the treatment of either R. aquatilis No-16 or SBC alone. Also R. aquatilis No-16 inhibited egg hatching even at 2% concentration of the bacterial suspension (1×10 CFU/ml). When cucumber seeds treated with this bacterial suspension at the concentration of 1×10 CFU/ml, a good amount of the bacterial population was recovered from the hypocotyls of the plant at the concentration of about 1×10 CFU/cm hypocotyl, indicating that this antagonistic bacterium was established and survived well on the plant. All of these results suggest that R. aquatilis No-16 may have a good potential to be developed as a biocontrol agent against the root-knot nematode on cucumber, possibly together with SBC as an additive.

Fusarium wilt of lettuce has caused a serious disease in a lettuce cultivation field of greenhouse during high temperature season. This pathogen is very difficult to control in a repeated lettuce cultivation field. A Strain of Bacillus subtilis GG95 antagonistic Fusarium oxysporum f. sp. lactucae was isolated from the lettuce cultivated soil. The antifungal activity by using filterpaper disc method showed inhibition zone on the radial growth F. oxysporum causing fusarium wilt. B. subtilis GG95 was cultured in a lab fermenter with TSB media (28 C, pH7) for 24hr. This strain was treated with 1×10 cfu/ml density, two times in a seedling tray(500ml/tray) and was sprayed three times with 10 cfu/ml density in a cultivating field greenhouse. After field spraying of B. subtilis GG95, the change of soil microflora was observed that density of Actinomycetes in a treated soil was increased 5 times compared to untreated plots. The control effect of GG95 treated plot showed 80%. The lettuce yield was increased to 17%, and sprout rate was 2% respectively higher than untreated plots.

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Han, Min Hye Jung, Young Jin Koh and Jae-Seoun Hur*. Korean

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Investigations of field performance of commercial biofungicides against powdery mildew of cucumber. Young Jin Koh , Gyoung 1

Hee Kim , Kyoung Youn Jo , Ju Hoon Cha , Jae-Seoun Hur and Sung-Jun Hong . Department of Plant Medicine and 1Department of 1

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tural Research and Extension Service, Hwaseong 445-784, Korea

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Antifungal activity of endolichenic fungi isolated from Korean and Nepalese lichens against plant pathogenic fungi. Keon Seon Lichen Research Institute, Sunchon National University, Suncheon 540-742, Korea

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Environmental Education, Sunchon National University, Suncheon 540-742, Korea; Organic Agriculture Division, National Academy of Agricultural Science, Suwon 441-707, Korea 2

Powdery mildew is one of the most destructive diseases on cucumber. Various kinds of chemical fungicides have been commonly used to control the disease. Recently, use of biofungicides as a substitute for chemical fungicides is increasing at farmers greenhouses, but their control efficacies is still controvertible. Six commercial biofungicides were investigated for their control efficacies when sprayed 3 to 5 times at 7-day-intervals from the early infection stage of powdery mildew on cucumber at the naturally infected greenhouse to develop their field performance enhancement techniques. One of the biofungicides applied 3 times exhibited more than 50% of control efficacy, but the other ones exhibited less than 50%. However, their control efficacies gradually increased in proportion to spray times. Four biofungicides applied 5 times resulted in more than 50% of control efficacies. Mixture of two different kinds of biofungicides enhanced their control efficacies more than single application of any of the biofungicides. However, the control efficacies of the biofungicides were

Endolichen fungi are a group of unexploited fungi inhabiting inside of lichen thalli. Antifungal activity of lichen endophytes (endolichenic fungi: ELF) against plant pathogenic fungi were investigated to develop novel biofungicides. Isolation of ELF was attempted from lichen thalli of 41 species collected in Jeju island, Korea and those of 90 species collected in Nepal with use of the surface sterilization method. Among 414 ELF strains isolated from Jeju Island, 34 representative isolates of various morphotypes were selected for investigation. Thirteen isolates from Nepal were also assessed. Dual culture bioassay was used for evaluation of antifungal activity against Alter-

naria alternate, Botrytis cinerea, Colletotricum acutatum, Fusarium oxysporum, Magnaporthe grisea, and Rhizoctonia solani. Among the

34 isolates of Jeju island, 9 ELF effectively inhibited more than 40% of mycelial growth of the pathogens, compared to control. Eight isolates from the tested Nepalese EFL also showed strong antifungal activity against the pathogens. Especially, three Nepalese ELF of NP141-2, NW151-2-2 and NP20-4 isolated from crustose lichen, Everniastrum nepalense and Usnea eumitrioides, respectively, produced red-colored pigment into PDA and exhibited complete inhibition of mycelial growth of R. solani. In general, C. acutatum and B. cinerea seemed to be more sensitive to the ELF than other pathogens.

Abstract of Presentations On the other hands, there was no significant activity of the ELF against F. oxysporum. Phylogenetic analysis of ITS sequences showed that the antifungal ELF were much closer to the known endophytic fungi than lichen-forming fungi. This suggested that endolichenic fungi could be related with fungal components associated with host tree bark. The study demonstrates that there are diverse endolichenic fungi inhabiting in lichen thalli and can be used as unexploited bioresources to develop biofungicides based on natural substances.

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Submerged air-lifting bioreactor for mass production of antifungal lichen substances. Jung A Kim, Young Jin Koh and Jae-Seoun

Hur*. Korean Lichen Research Institute, Sunchon National University, Suncheon 540-742, Korea

Lichen-forming fungi (LFF) produce unique antifungal substances, which can be used as leading compounds for novel biofungicdes. Submerged air-lifting bioreactor was developed as a novel cultural system for mass production of lichen substances. The bioreactor was designed to stimulate mycelial growth of LFF and produce lichen substances in a large scale. Mycelial growth of Cladonia macilenta increased by 2.3 times high in the bioreactor, compared with conventional method of shaking culture. Production of biruloquinone, purple-colored lichen substance of the fungus, was also increased by 2.6 times high in the bioreactor than that in shaking culture. The results suggest that mass production of lichen substances can be feasible for industrial application by culturing lichen-forming fungi with use of submerged air-lifting bioreactor.

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sequencing of internal transcribed spacer region. Yun Jung Go , So Young Kang , Sang-Woo Lee and Yong Chull Jeun . Faculty of 1

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Koh and Jae-Seoun Hur*. Korean Lichen Research Institute, Sunchon National University, Suncheon 540-742, Korea

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Bioscience and Industry, College of Applied Life Sciences, Jeju National University; Gyeonggi Province Agricultural Research and Extension Services, Hwaseong 445-972, Korea 2

The bacterial isolates from Jeju Island and one bacterial isolates from Jeonnam province were identified by analysis of the ribosomal DNA sequence in the internal transcribed spacer (ITS) region. Among total 14 bacterial isolates tested, 7 bacterial isolates showed either antagonistic against plant pathogens or induced systemic resistance against plant diseases and 4 bacterial isolates were showing both effects. The others 3 bacterial isolates did not show neither of them. Using universal primers, all of the bacterial DNA samples were amplified. Among the effective bacterial isolates identified, Bacillus spp. were the most popular bacteria among the isolates i.e. 4 isolates were identified as B. cereus and the others 2 isolates as B. weihenstephanensis and B. circulans. The bacterial isolate CHT2008 from Jeonnam was also identified as B. cereus. Two isolates were identified as Burkholderia gladioli. And the others 3 isolates as Pseudomonas putida, Miamiensis avidus and Acinetobacter quenomosp. On the other hand, among 3 non-effective bacterial isolates, two isolates were identified as Acinetobacter quenomosp and one as Renibacterium salmoninarum. The clade gramme showed that the B. cereus KRY505-3, MRL412 and CHT2008 were genetically similar whereas B. cereus TRL2-3 was a little bit different from the strains.

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Resistance induction in potato tissue culture by pre- or post-inoculation with Burkholderia gladioli TRK2-2 against late blight. So

Young Kang , Chan Woo Kim , Ki Deok Kim and Yong Chull Jeun . 1

Herbicidal activity of lichen substances extracted from lichen thalli and lichen-forming fungi. Jung A Kim, Heng Luo, Young Jin

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Faculty of Bioscience and Industry, College of Applied Life Sciences, Jeju National University, Ara-dong, Jeju-si, Jeju 690-756, Korea; Division of Biotechnology, College of Life Sciences and Biotechnology, Korea University, Anam-dong, Sungbuk-ku, Seoul 136-713, Korea 1

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Lichen substances are unique compounds having several bioactivities, such as antitumour, antibacterial, antifungal, antiviral, antiinflammatory and antioxidant activities. In this study, herbicidal activity of unexploited lichen bioresources was screened to develop novel bioherbicides with use of the thalli extracts of 16 lichen species collected from extreme environments such as highland and Antarctic and the mycelial extracts of more than 300 lichen-forming fungi (LFF) deposited at Korean Lichen Research Institute, Sunchon National University. LFF extracts of Graphis sp., Megalospora tuberculosa, Menegazzia sp., Physcia sp., Ramalina terebrata, and Rhizoplaca sp. inhibited seed germination and caused foliar chlorosis of Arabidopsis in a dose-dependant manner. Thalli extracts of Cladonia furcata (IC 167µg/ml), Cetraria aculeata (IC50 115µg/ml), Cetraria islandica (IC50 140µg/ml), Lethariella zahlbruckneri (IC 112µg/ml) and Letharia vulpina (IC50 24µg/ml) exhibited strong herbicidal activity. Vulpinic acid was found to be the herbicidal constitute of L. vulpine. The results suggested that lichen substances have a potential of novel bioresources to develop bioherbicides based on natural substances.

Late blight is one of the destructive diseases causing yield loss in not only seed potato but also field potato production. For supplying virus free tubers, most seed potato is produced by tissue culture, in which application of commercial pesticides is limited. However, there are only a few bio-fungicides effective against late blight in tissue culture. In our previous study, a bacterial isolate TRK2-2 was selected as a resistance inducing agent by a greenhouse experiment. In this study it was investigated the efficacy of the bacterial isolate TRK2-2 against late blight in tissue culture. Pre-inoculation of the bacterial suspension caused suppression of disease severity of late blight whereas postinoculation did not. Pre- or post treatments of DL-3-amino butyric acid, a well known resistance inducer, were showed similar results with those of TRK2-2. On the other hand, using analysis of ribosomal DNA in the internal transcribed spacer, the bacterial isolate was identified as Burkholderia gladioli. Based on these results, it was suggested the Burkholderia gladioli TRK2-2 may be useful in the potato tissue culture as a resistance induction agent.

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Characteristics of effective bacterial isolates from Jeju Island by

ACC deaminase producing strain ST45 confers resistance to high

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salt stress in plant. Hyeon Seong Yoon, Jinhee Choi, Jae-Chan Lee, Dong-Jin Park and Chang-Jin Kim. Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 305-333, Korea

Ethylene is a gaseous phytohormone that plays important role in many developmental process and environmental fitness responses of plant. Ethylene biosynthesis is well studied by biochemical and genetic approaches. ACC (1-aminocyclopropane-1-carboxylate) is precursor of ethylene and it is hydrolyzed by ACC deaminase into αketobutyrate and ammonia, and thereby lowering the level of ethylene in the plant. Decreased ethylene levels make the plant to have more resistant to environmental stresses. We screened ACC-deaminasecontaining bacteria from soil and selected ST45 showing high ACCdeaminase activity among 1200 isolates. The result of through 16s rDNA sequencing and fatty acid analysis indicated that ST45 belonged to the family Enterobacteriaceae. Under high salinity stress condition (210mM NaCl), ST45 promoted plant growth, such as root growth, shoot growth, and fresh weights. These results suggest that the ST45 has PGPB (Plant growth-promoting bacteria) effect by the alleviation of various stresses condition such as salinity on plant growth.

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Control of bacterial wilt of tomato plants by Pseudomonas putida LSW17S under different nutrient conditions. Sang-Woo Lee *,

wilt could be effectively controlled by LSW17S treatment in low level of P concentration due to accumulation of high SA concentration in tomato plant.

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Fermentation and Formulation of Acremonium strictum BCP.

Bong-Su Kim , Taek-Su Shin , Jae-Sung Im , Seong-Gyun Woo , JinCheol Kim , Gyung-Ja Choi , Jae-Ho Lee . GreenBiotech Co. Ltd., 1

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Gyoha-eup, Paju, Gyeonggi-do 413-830, Korea; Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea 2

Acremonium strictum BCP is the hyperparasitic fungi on gray mold disease, caused by Botrytis cinerea. Weve researched the optimal medium and the large scale fermentation method of A. strictum BCP

to develop commercial biopesticide. To design the optimal medium for large scale fermentation, optimal nitrogen and carbon source was examined. After this, large scale fermentation process (temperature, pH, fermenting period, initial inoculums size, agitation rate, aeration rate) was controlled in jar fermenter(5L) and large scale fermenter (500L, 5000L) condition. And weve formulated A. strictum BCP as powder shape to improve spore stability and efficacy. Formulated trial products were treated on tomato greenhouse. Efficacy on tomato gray mold was more than 80%, and no harmful effect was shown.

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Sang-Yeon Sim , Su-Yeon Lee , Myeon-Whoon Seo , Jae Wook Lim and Yong-Hwan Lee . Gyeonggi-Do Agricultural Research and 1

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Extension Services, Hwaseong 445-300, Korea; Department of Agricultural Biotechnology and Center for Fungal Genetic Resource, Seoul National University, Seoul 151-921, Korea 2

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Control of Scab of Japanese Apricot by environment- friendly spraying program. Gil-ho Shin , Yang Gi Na, Jong Bun Seo, Hyo 1

Jin Jung, Bong Gi Yun, Jong Sup Shin , Young Jin Ko and Kyung Yeon Jo. Jeonnam Agricultural Research and Extension Service, 2

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Naju 520-715, Korea; Suncheon-Si Agricultural Technology and Extension Center, Suncheon 540-742, Korea; Department of Plant Medicine, Suncheon National University, Suncheon 540-742, Korea 2

Bacterial wilt caused by Ralstonia solanacearum is a disease widely distributed in the world. It takes place in plant root and spreads through the vascular system, causing plant pith to decay into a slimy mass. If it infects and establishes on plant, it is not easy to control of this bacterial wilt disease. Pseudomonas putida LSW17S (LSW17S) isolated from tomato root decreased the disease severity of bacterial wilt caused by Ralstonia solanacearum. Treatment of LSW17S decreased severity of bacterial wilt in cultivar Housemomotaro which is susceptible to R. solanacearum and its control value was 56% in pot test. When tomato plants were cultivated in the normal nutrient solutions, there is no significant difference in plant growth. However, when plants were cultivated in nitrogen and calcium deficient nutrient solution, leaf width and fresh weight were decreased by bacterial treatment. On the contrary, growth elements of tomato plants grown in phosphorus deficient nutrient solution were significantly promoted by LSW17S treatment as compared to those of control. On the other hand, P starvation highly increased population density of LSW17S and SA concentration than other treatments. Furthermore, bacterial treatment increased phosphorus and iron absorption in the phosphors limited condition. In different P concentration treatment, LSW17S effectively protected bacterial wilt in a low P nutrient concentration. Disease severity was decreased, as P concentration was decreased. Moreover, in a low P concentration, SA concentration in leaf tissues was higher than in high P concentration and population density of LSW17S was also higher in low P condition. Moreover, LSW17S effectively increased P uptakes in 0 ppm or 5 ppm P nutrient conditions, and Fe uptake in all nutrients. These results indicate that tomato

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Scab caused by Cladosporium carpophilum are the most common and serious diseases of Japanese apricot in the environment-friendly farmers' orchards. The effect of environment-friendly products against C. carpophilum through in vitro and in vivo test was examined. Application environment-friendly products(EFPs), Shooting Star®, Cool®, Doyeolee®, and NaDCC reduced in vitro growth of the pathogen. To develop a spraying program during growth stages of Japanese apricot, environment-friendly spraying program(EFSP), applied EFPs 5 times and low pesticide spraying program(LPSP), applied pesticides 3 times, were applied to three experimental plots. The treatments resulted in 75.2% and 84.1% control efficacies in EFSP and LPSP, respectively against scab of Japanese apricot in the field. Control value was not significantly different both the spraying programs in the field. Disease incidence of the control was 19.9%.

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Biological control of soft rot on Chinese cabbage using Lactobacillus, Lactococcus, and Paenibacillus under greenhouse and field conditions. Anupama Shrestha , Eun Chang Kim , Duck Hwan 1

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Park , Jang Hyun Hur and Chun Keun Lim . Laboratory of Bacterial Genetics and Biotechnology, Division of Bio-Resources Technology, College of Agriculture and life Sciences, Kangwon National University, Chuncheon 200-701, Korea; Division of Biological Envi1

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Abstract of Presentations ronment, College of Agricultural and Life Sciences, Kangwon National University, Chuncheon 200-701, Korea

Bacterial soft rot caused by Erwinia carotovora subsp. carotovora occurs worldwide and is one of the most hazardous pathogen which damages Chinese cabbage production in Korea. Three bio-control strains (BCAs) Lactobacillus strain KLF01 Lactococcus strain KLC02 and Paenibacillus strain KPB3 were screened to see their in vitro antagonistic effect against bacterial soft rot caused by Erwinia carotovora subsp. carotovora. We also conducted under greenhouse and field conditions to evaluate the control effects of BCAs against soft rot. At results, all three strains were able to show clear zone against plated soft rot pathogen. For greenhouse test, BCAs (1 ×10 CFU/ml) were applied as a drench to the seedlings and then E. carotovora subsp. carotovora (1×10 CFU/ml) grown on Mannitol Glutamate Yeast (MGY) broth with mineral oil in the ratio 4:1 was inoculated in the soil treated with BCAs 5 day post inoculation (dpi) in completely randomized experiment by triplicate. Disease progress assessment was done every 4 , 8 and 12 dpi. Bio-control efficacy was more significantly effective in the treatments by Lactococcus sp. and Paenibacillus sp. as 64, 50, 56% and 66, 57, 58% according to date of evaluation. In case of Lactobacillus sp., control effect was relatively lower than treatments of Lactococcus sp. and Paenibaciilus sp., but its efficacy was still significantly observed when compared to negative control (only water treatment). Furthermore, Lactobacillus sp., Lactococcus sp., and Paenibacillus sp. showed 55%, 60% and 62% bio-control efficacy respectively in field test that was conducted from early March to late July of 2009 at experimental farm of the Kangnan National University. Thus, we suggest that these strains can be useful as a bio-control agent against soft rot caused by Erwinia carotovora subsp. carotovora. 8

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Antifungal activity of chaetoglobosin A from culture filtrate of Chaetomium globosum CNU 9051 against plant pathogenic fungi.

Eun Young Seo , Yang Kyu Kim , Bong Sik Yun , and Seung Hun Yu . Department of Applied Biology, College of Agriculture and Life 1

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Sciences, Chungnam National University, Daejon 305-764, Korea; Biotechnol., Coll. Environ. Bioresour. Sci., Chonbuk National Univ., Chonju 561-756, Korea 2

In our previous studies, we selected antagonistic fungal isolate CNU 9051 among the endophytic fungi from tomato root. The fungus was identified as Chaetomium globosum based on morphological characteristics and rDNA ITS sequence analysis. An antifungal substance was isolated from the culture filtrate of CNU 9051. It was extracted by chloroform and purified by silicagel column chromatography and TLC analysis. The molecular formula of the isolate was established by NMR and HPLC analyses and identified as chaetoglobosin A (C H N O ). The antifungal substance strongly inhibited the mycelial growth of Alternaria panax, Botrytis cinerea and Sclerotinia sclerotiorum. It also inhibited mycelial growth of Colletotrichum acutatum, Fusarium oxysporum and Phytophthora capsici and also inhibited conidial germination of B. cinerea. 32

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Control Effect of Commercial Biofungicides for Control of Pow-

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dery Mildew on Tomato in Organic farm. Ji Hee Han, Hyun Young

Shin, Kyung Seok Park, Sang Yeop Lee and Jin Woo Park. Microbial Resources Lab., Agricultural Microbiology Division, National Academy of Agricultural Science (NAAS), RDA, Suwon 441-707, Korea

Tomato powdery mildew is caused by Golovinomyces cichoracearum. This pathogen decreases yield and quality of agricultural

product by reducing of chlorophyll in the leaves of plants. There are some chemical fungicides for tomato powdery mildew, but there is no biological fungicide that is officially registered. We began the experiment in February 19th at organic tomato farm in Gongju. Variety of tomato is doterang dia which is planted in the greenhouse on December 24, 2008. To screen effective biofungicide for tomato powdery mildew, three commercial biofungicides (A: Paenibacillus polymyxa, B: Bacillus subtilis, Q-fect WP : Ampelomyces quisqualis AQ94013) are experimented in the greenhouse. Three biofungicides were sprayed three times at seven-day interval at the begging stage of the disease, and investigated the disease incidence of powdery mildew during four weeks. The disease incidence of control plot was increased from 10% to 25% during experiment period. The control effect of A, B and Q-fect WP on tomato powdery mildew was 2.5%, 7.1% and 86.9%, respectively. Therefore, this experiment suggested that Q-fect WP was effective to control for tomato powdery mildew.

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Antagonistic activity of endophytic bacterium isolated from chili pepper (Capsicum annum L.) plant. Narayan Chandra Paul, Eun

Young Seo, Li Meijia and Seung Hun Yu. Department of Agricultural Biology, Chungnam National University, Daejeon 305-764, Korea

Endophytic bacteria were isolated from the surface sterilized tissues of leaves, stems and roots of chili pepper plants (Capsicum annum L.) grown in the field in Korea. One hundred fifty isolates were evaluated for the antifungal activity against three major fungal pathogens (Phytophthora capsici, Colletotrichum acutatum and Fusarium oxysporum) of chili pepper in vitro. Twenty isolates showed antifungal activity against at least one pathogenic fungus of chili pepper. The isolate CNU 082142 exhibited the maximum ability to reduce the Phytophthora capsici growth in dual culture assay. The isolate was further screened for the reduction of disease severity caused by P. capsici using detached leaves and seedling assay and showed varying levels of antagonism against P. capsici in both detached leaves and seedlings assay. These results indicate that the in vivo seedling assay can be used as a rapid and more accurate technique for the selection of promising biocontrol agents against P. capsici.

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Effect of Plant Extracts on Control of Rice Leaf Blast. Beom

Ryong Kang, Sug-Ju Ko, Do-Ik Kim, Seon-Gon Kim and Hong-Jae Kim. Environment-Friendly Agricultural Research Institute, Jeollanamdo Agricultural Research and Extension Services, Naju 520-715, Korea

More than 30 plant extracts tested for their antifungal activity against Magnaporthe grisea causing rice leaf blast. Analysis of the recovered material was done by rotary evaporator. The components of methanol and ethanol on heating extraction showed nearly quantitative recov-

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ery after 14 h(more than 95%), but soaking required 40 h to reach 80% recovery. Methanol was shown to be an excellent extracting solvent for most of the plant part. In this experiment methanol, ethanol, acetic acid, hexane and ethylacetate extracts inhibited the radial growth of the fungi using impregnated paper disc methods. In a test with M. grisea, strong activity was obtained from the extracts of Anemarrhena asphodeloides, Albizzia julibrissin, Coptis chinensis, Rhus chinensis, Glycyrrhiza uralensis, Schizandra chinensis, and Aconitum ciliare for 72 h. Extracts of Coptis chinensis roots, Anemarrhena asphodeloides roots, Angelica gigas roots, and Albizzia julibrissin

barks showed potent fungicidal activity against the pathogens when treated with 100 diluted concentrations in greenhouse. Coptis chinensis roots extract displayed control values of more than 65% and more than 55% mixed Rhus chinensis and Angelica gigas against rice leaf blast in the field. These results suggest that Coptis chinensis- and Angelica gigas-derived materials might be useful for developing new type fungicides, or as natural fungicidal extracts against rice blast.

semi-selective medium and their growth inhibitory ability of the several fungal pathogens of grape were assayed. Antagonistic isolates against the grape fungal pathogens were selected on growth inhibition assay on agar. Then, growth inhibition ability of the selected antagonistic isolates were tested using the culture filtrate. One of the isolates, FM16 inhibited the growth of all fungal pathogens of grape including the ripe rot pathogen, Glomerella cingulata. Microscopic observation, the cellular fatty acid profile by MIDI system and 28S rDNA sequence indicated that the FM16 is Candida sp. The spray of FM16 cell suspension reduced the ripe rot significantly in field trial. Control value of the FM16 suspension was equivalent to the dithianon WP's, which is the registered fungicide for the ripe rot of grape.

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Antiviral Activity of Pseudomonas oleovorans and Acinetobacter antiviralis sp. Nov. against Vector Transmission of Rice Stripe Virus. Minho Lee , Yurim Jeong , Nam Gyu Kim , Cheol Nam Park , Jong Gwan Kim , In Cheon Hwang , Tae Yu Yun , Beom Seok Kim , Ki Hyun Ryu and Mun Il Ryoo . College of Life Science & 1,2

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Biological control of red pepper anthracnose and cucumber powdery mildew by Bacillus amyloliquefaciens KB-JKM. Jae Jun-

Lee*, Myoung Heun-Nam, Keun Hwan-Lim, Hyun Su-Lee, Jae PilChoi and Young Gyun-Kim. Institute of Biotechnology, Kobio Co. Ltd

A strain Bacillus amyloliquefaciens KB-JKM was isolated from the tomato soil at Hwasung Korea. In the course of screening for antifungal activity against nine plant pathogenic fungi (Alternaria kikudes, Botrytis sp., Collectorichum coccodes, Fusarium oxysporum, Phytopthora capsici, Pythium sp., Cladosporium fulvum, Sclerotinia sp., Rhizoctonia solani) of isolate, strain KB-JKM showed strong antibiotic activity against all pathogenic fungi. In determining its relationship on the basis of 16S rDNA sequence, KB-JKM strain was most closely related to Bacillus amyloliquefaciens (AY603658) (99.84%). KB-JKM WP (2.1×10 cfu/g) was produced from culture broth B. amyloliquefaciens KB-JKM. We have discovered that the stability of antifungal compounds in powder compositions after 6 months. Both of treatment KB-JKM WP (2.1×10 cfu/g) 250 and 500 times diluents showed excellent antagonistic effect against anthracnose of red pepper, Specially the treatment of 250 times showed control value of 84.4%. These results show that KB-JKM can be used as a biofungicide for the control of anthracnose (Colletotrichum coccodes) and cucumber powdery mildew (Sphaerotheca fuliginea) on red pepper and cucumber plants. T

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C-46

Candida sp. FM16 Reduces Severity of the Ripe Rot of Grape. Jung-Soon Park , Mi-Soon Baek , Seung-Up Yang , Jong-Hwan Park and Jae-Soon Cha . Department of Plant Medicine, Chungbuk 1

2

1

1

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1

National University, Cheongju, 361-763, Korea; National Institute of Horticultural and Herbal Science 475th, Imk-dong Jangan-gu Suwon, 440-706, Korea 2

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3

3

4

1

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Biotechnology, Korea University, Seoul 136-713, Korea; Division of Applied Entomology, National Academy of Agricultural Science, Suwon 441-707, Korea; Central Research Institute, Kyung Nong Co., Ltd., GyeongJu 780-110, Korea; Division of Environmental & Life Sciences, Seoul Women’s University, Seoul 139-774, Korea 2

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4

Two biochemicals, KNF2016 and 2022 derived from Pseudomonas oleovorans and Acinetobacter antiviralis sp. Nov., showing high antiviral activity to several filamentous plant viruses such as TMV, PMMoV and CGMMV, have been ready to be registered for biochemical pesticide by Kyung-Nong Co., Ltd.. Rice stripe virus (RSV) is also filamentous, and damages rice seriously by vector transmission of small brown planthopper, Laodelphax striatellus, in the East Asia. Thus, we have assayed the antiviral activity of the strains against RSV. The microbial formulations were sprayed on rice plants, and the RSV-viruliferous vector insects were infested on the rice plants 1, 3, or 5 days after treatment. As the result, the applications showed significantly high antiviral activity against RSV transmitted by the vector insect after treatment. Infectivity of the virus was decreased over 30% in the treatments of 250-/500-dilluted solutions. Thus, resistance of test rice plant, which is originally „high susceptive cultivar, was revealed as „high resistant or „resistant in the treatments. The antiviral activity against RSV infection was decreased according to the increase of time (day) after treatment. However, molecular diagnosis showed that the viral transmission was able to be inhibited over 2 weeks after treatment on rice plant infested with the RSV-viruliferous vector insects.

C-48

Identification and Selection of Paenibacillus polymyxa KC028 as potential biological control of several plant disease. Jiae Seo , 1

Seakwon Choi , Dongkyu Ko , Jiyoung Park and YoungKeun Yi . 1

2

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B&L Agro, Andong, Gyeongbok 760-380, Korea; Gyeongbuk Institute for Bio industry, Andong, Gyeongbok 760-380, Korea; College of Natural Sciences, Andong National University, Andong, Gyeongbok 760-740, Korea 1

2

3

Saprophytic yeasts have been studied as potential biological control agents and biological control yeasts have been used to manage postharvest losses due to fungal pathogens in many fruit crops including grapes. In this study, yeasts were isolated from the grape using the

To select the new antifungal bacteria, fifty of antifungal bacteria were

Abstract of Presentations isolated from various environments located in Gyungbuk area. From these isolated antifungal bacteria KC028 was selected and was identified as Paenibacillus polymyxa by biochemical and morphlogical analyses. The optical growth conditions were 35 C for temperature and NA+PDA for method. It had potential biological control of several plant disease in indoor experiments (Gibberella fujikuroi, Cero

cospora kikuchii, Phytophthora capsici, Colletotrichum acutatum, Colletotrichum gloeosporioides, Pyricularia grisea, Glomerella glycines, Rhizoctonia solani, Botryosphaeria dothidea, Fusarium oxysporum f. sp. cucumerinum). This study was carried out with the

support of ‘GIB Bio project-01’ provided by Gyeongbuk Institute for Bio industry.

C-49

Analysis of fusaricidins produced by Paenibacillus polymyxa E681 and its possible role in a bacterial determinant of induced systemic resistance on pepper. Seong-Bin Kim, Jung Wook Yang, Choong-Min Ryu and Seung-Hwan Park. Industrial Biotechnology & Bioenergy Research Center, KRIBB, Daejeon 305-806, Korea

Fusaricidins are group of lipopeptide antibiotics produced by Paenibacillus polymyxa and composed of six amino acids residues in addition to 15-guanidino-3-hydroxypentadecanoic acid. Fusaricidins have an excellent antifungal activity against plant pathogenic fungi and gram-positive bacteria. We reported previously the complete sequence of fusaricidin synthetase gene (fus A) identified from the genome sequence of a rhizobacterium, P. polymyxa E681. In this study, the fusaricidins were purified from culture broth of strain E681 by butanol extraction, silica gel and sephadex LH-20 column chromatography. Finally, six different fractions having molecular masses of 883, 897(2), 911(2) and 925 Dalton were obtained by reverse phase high performance liquid chromatography as a fusaricidin group of cyclic depsipeptide. The amino acids composition of the fusaricidins have been analyzed and showed that those are fusaricidin A, fusaricidin B, LI-F5a, LI-F5b, LI-F8a and LI-F8b. Besides antifungal and antibacterial capacity, the fusaricidins were exploited to assess response to plants by examining elicitation of induced systemic resistance (ISR) against a bacterial pathogen in pepper. The purified fusaricidins drench-applied in the pepper seedlings and tested on ISR against Xanthomoans axonopodis pv. vesicatoria seven days after leaf-infiltration. Even 11 nM fusaricidin application significantly decreased symptom development compared to water control. Our results were confirmed the production of fusaricidins from P. polymyxa E681 that harbor a fusaricidin synthesis gene cluster and indicated its application on ISR on pepper.

C-50

Pepper growth promotion and biocontrol characteristics by phosphate solubilizing bacteria Chryseobacterium indologenes strain ISE14 against Phytophthora capsici. Mee Kyung Sang , Inn1

Shik Myung and Ki Deok Kim . Laboratory of Plant Disease and 2

1

1

Biocontrol, College of Life Sciences and Biotechnology, Korea University, Seoul 136-713, Korea; Division of Agricultural Microbiology, National Academy of Agricultural Science, RDA, Suwon 441707, Korea

286

this strain in relation with plant growth promotion and biocontrol activities against Phytophthora capsici on pepper. Strain ISE14 significantly reduced disease severity and areas under the disease progress curves of 5-week-old pepper plants inoculated with P. capsici compared to buffer-treated control. Strain ISE14 was identified as Chryseobacterium indologenes based on morphological, physiological, biochemical, Biolog, fatty acid methyl ester, and 16S rDNA sequence analyses. Strain ISE14 significantly promoted pepper growth compared to the buffer-treated control through production of acid phosphatase and solubilization of phosphate. This strain colonized pepper roots efficiently and significantly altered microbial activity in treated soil of pepper. Strain ISE14, identified as C. indologenes, is both an excellent plant growth promoting bacteria (PGPR) with phosphate solubilization activities and biocontrol agent against Phytophthora blight of pepper, protecting infection courts through effective root colonization by biofilm formation. Therefore, C. indologenes strain ISE14 could be used as a PGPR and biocontrol agent for controlling Phytophthora blight of pepper.

C-51

Effects of compost water extracts on biological control and inducted systemic resistance against Phytophthora blight of pepper. Mee Kyung Sang , Eui Nam Kim , Jeong-Gyu Kim and Ki 1

1

2

Deok Kim *. Laboratory of Plant Disease and Biocontrol, College 1

1

of Life Sciences and Biotechnology, Korea University, Seoul 136-713, Korea; Division of Environmental Science and Ecological Engineering, College of Life Sciences and Biotechnology, Korea University, Seoul 136-713, Korea 2

In this study, we investigated water extracts of composts from commercial fertilizer plants for use of plant production on controlling root and foliar diseases by Phytophthora capsici on pepper plants. Water extracts from composts (CWE) Iljuk-3, Iljuk-7, Shinong-8, and Shinong-9 significantly (P=0.05) inhibited zoospore germination, germ tube and mycelial growth of P. capsici and reduced pathogen population, and pepper root and foliar infections by P. capsici in pepper plants. However, P. capsici germination, and disease incidence and severity were not different with treatments of autoclaved or filtered, and raw (unsterilized) CWE. In addition, CWE significantly (P=0.05) suppressed the pathogen infection in leaves through induced systemic resistance when CWE applied into roots of pepper plants. The tested CWE could enhance expression of PR genes and enzyme activities including â-1,3-glucanase, chitinase, and peroxidase, resulting in enhanced plant defense against P. capsici infection in pepper plants. Moreover, the CWE enhanced chemical and structural defenses including hydrogen peroxide production in leaves and lignin accumulation in stems of pepper. From these results, it has been suggested that a heat-stable chemical(s) of tested CWE could suppress root and foliar infections by P. capsici in pepper plants. These suppressions might result from a direct inhibition of P. capsici development and rhizosphere population, and an indirect inhibition of foliar infection through ISR.

2

This study aims to identify bacterial strain ISE14 and to characterize

C-52

Enhanced Induce Systemic Resistance against Cucumber anthracnose pathogen by Bacillus subtilis strain B4 that grown on different culture media. Kyungseok Park*, Jae Eun Lee, Ji Hee Han

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and Jin Woo Park and Sang Yeob Lee. Division of Agricultural Microbiology, National Academy of Agricultural Science, RDA Suwon 441-707 Korea

One of selected plant growth promoting rhizobacterial strain, Bacillus subtilis B4 played an important role in Induce systemic resistance (ISR) and plant growth promotion in various plants. After B. subtilis B4 was incubated on KB, TSA, NA and LBA respectively, each bacterial cell suspension were drenched on cucumber rhizosphere soil or infiltrated on cucumber leaf, the plants were showing different activity of Induced systemic resistance as well as antifungal activities against major plant pathogens. B4 that grown on Nutrient Broth(NB) and Minimal salt(MS) showed enhanced systemic resistance against Colletotrichum orbiculale than Tryptic soy broth(TSB) and Luria broth(LB) on cucumber plant. Whereas, the treatment of TSB and LB growing B4 showed stronger antifungal activity against Fusarium oxsporum, Phytophtora capsici, and Rhizoctonia solani than NB and MS media. In this study, the results indicate that selection of suitable media among culturing media is significantly important in the test of induced systemic resistance.

C-53

Enhancement of antifungal activities against Phytophthora capsici by heat treatment of culture filtrates of Bacillus subtilis B4. Kyungseok Park*, Yeong Eun Cho, Sang Min Cho, Kyung Soo Park and Jin Woo Park. Division of Agricultural Microbiology, National Academy of Agricultural Science, RDA Suwon 441-707, Korea

Selected strain of the plant growth promoting rhizobacteria, Bacillus subtilis B4 has been reported to induced systemic resistance in redpepper plant as well as showing strong antifungal activity against Phytophthora capsici In Vitro and In Vibo. The MeOH fractions (Nos. 0, 10, 20, 40, 60, 80, 100 fractions) of B4 cell culture collected through C-18 chromatography was tested for antibiosis against major fungal pathogen, Phytophthora capsici. Among them, fraction numbers 80 and 100 showed significant growth suppression of fungal mycelium. Moreover, 0, 10, 40 and 100 fractions dissolved in H2O and MeOH solvents were found to be increased GUS gene expression which related to PR1a in tobacco compared to control plants. In this study, we have emphasized that the heat treatment of B. subtilis B4 culture filtrate can enhance the level of antibiosis and disease resistance. Likewise, it helps to increase the level of metabolites production by B4 strain.

D-1

Mechanism of Action and Selectivity of Pyribencarb: The New Type of Fungicide in Qo Inhibitors. Satoshi Kataoka, Makiichi Takagaki, Koichiro Kaku and Tsutomu Shimizu. Life Science Research

its mode of action and selectivity. Pyribencarb potently inhibited succinate-cytochrome c reductase (SCR) activities of cucumber gray mold (Botrytis cinerea), leaf spot (Corynespora cassiicola) and decylubiquinol-cytochrome c reductase (UCR) activity of B. cinerea. Pyribencarb inhibited the UCR of B. cinerea in the uncompetitive manner with respect to decylubiquinol as same as strobilurin fungicides, and the substrate-dependent inhibition constant was found from calculation to be 13 nM. These results suggested that the target site of pyribencarb is cytochrome b of the complex III in the electron transport system of the respiratory chain. Also, SCRs of the resistant strains of B. cinerea and C. cassiicola against the strobilurin fungicide were highly resistant to strobilurin fungicides (kresoxim-methyl and azoxystrobin), but less resistant to pyribencarb, resulting in that the RS ratios of pyribencarb for the inhibitions of SCR activities of B. cinerea and C. cassiicola were smaller than those of strobilurin fungicides. On the other hand, inhibitory potency of pyribencarb to SCR activities of plants (tomato, kidney bean and cucumber), rat and carp was relatively weak compared with those QoIs. These results indicated that pyribencarb was a novel inhibitor of the complex III was superior to the well-known QoIs in terms of the selectivity and selective toxicity. The binding site of pyribencarb on cytochrome b was assumed to be a little different from those of strobilurin fungicides through comparison of amino acid sequences of plants, rat, carp and fungi.

D-2

Antifungal activity of pentaene microlide strevertene produced by Streptomyces sp. KP1404. Jeong Do Kim, Jae Woo Han, Eun

Young Kim, Sin Ae Kang, Hye Yoon Kim and Beom Seok Kim. Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University, Seoul 136-713, Korea

Necessity of novel fungicides useful for plant disease control is continuously growing due to environmental concerns and the appearance of fungicide-resistant strains of plant pathogen. Streptomyces sp. KP1404 which was isolated from soil sample collected in Guri, Korea showed antifungal activity against a variety of plant pathogenic fungi. The antibiotic substance ST-1 produced by Streptomyces sp. KP1404 was purified using various chromatographic methods. Structure information of ST-1 was obtained from spectrometric analysis using MS and NMR. The antibiotic compound isolated from KP1404 turned out to be strevertene which has a characteristic large lactone ring with pentaene moiety. In vitro bioassay, strevertene exhibited strong inhibitory activity on mycelia growth of plant pathogenic fungi such as Colletotrichum orbiculare, Diaporthe citri, Fusarium oxysporum f. sp. lycopersici. Magnaporthe grisea, Phytophtora capsici, Rhizoctonia solani.

Institute, Kumiai Chemical Industry Co., Ltd., Kikugawa, Shizuoka 439-0031, Japan

D-3

Pyribencarb is a novel benzylcarbamate type fungicide, which has jointly been developed by Kumiai Chemical Industry Co., Ltd. and Ihara Chemical Industry Co., Ltd. This fungicide has been proven to be active for a wide range of plant pathogenic fungi. In this presentation, potential inhibitory effects of this fungicide on the electron transport system of fungi, plants, rat and carp were examined to elucidate

Lee, Eun Kyung Jung, Zhang Yan, Cheol Soo Yoon and Byung Sup Kim. Department of Plant Science, Gangneung-Wonju National Uni-

Influence of application method on the efficacy of amisulbrom for control of clubroot of Chinese cabbage. Jae Eun Lee, Sang Jun versity, Gangneung 210-702, Korea

The ability of amisulbrom to control Chinese cabbage clubroot caused by Plasmodiophora brassicae was evaluated in field trials

Abstract of Presentations conducted in 2008-2009. Field trials were designed to develop and evaluate improved methods of application to control the disease. Experiment was accomplished in field located in Gangneungshi alpine area contaminated by Plasmodiophora brassicae. Application methods of amisulbrom used field trials were soil mixing, soil drenching, seedling socking, soil mixing+soil drenching, soil mixing+seedling socking, and untreated control before transplanting. As soil mixing of amisulbrom DP and soil drenching of amisulbrom WP provided 78.6% and 73.2% in control value in 2008 and soil mixing of amisulbrom DP and soil drenching of amisulbrom SC provided 96.4% and 93.6% in control value in 2009, this fungicide showed above or similar efficacy to standard fungicides. Among application methods, soil mixing+soil drenching before transplanting was the most effective for control of clubroot as 96.5% in 2008 and 97.2% in 2009, respectively. When yield of each treatment was examined, soil drenching, soil mixing+soil drenching and soil mixing provided 8.4%, 7.9% and 7.3%, respectively in increasing rate in fresh weight of above-ground part comparing with untreated control.

D-4

Occurrence of strobilurin-resistant isolates of Pseudocercospora vitis, causal fungus of grapevine leaf blight in Japan. Koji Inoue , 1

Natasya Koya and Akira Kawaguchi . Agricultural Experiment Station, Okayama Prefectural General Agriculture Center, Akaiwa-city, Okayama, 709-0801, Japan; Nagoya Plant Protection Station, The Ministry of Agriculture, Forestry and Fisheries of Japan, Nagoyacity, Aichi 455-0032, Japan 2

1

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2

The strobilurin fungicides, kresoxim-methyl and azoxystrobin, have been used to control ripe rot, downy mildew and leaf blight of grapevine for more than ten years in Okayama Prefecture, Japan. Failing efficacy of strobilurin fungicides to leaf blight was lately observed in some vineyards. Seventy seven isolates of Pseudocercospora vitis collected from ten vineyards in Okayama Prefecture were tested for sensitivity to a strobilurin fungicide in 2006. The MIC of azoxystrobin against mycelial growth in presence of 5mM n-propyl gallate was more than 400ppm in 61 isolates. A single point mutation (G143A) in the cytochrome b gene was found in the resistant isolates tested. Control efficacy of azoxystrobin against resistant isolates was very low in an inoculation test using pottedgrapevine plants. Most of these strobilurin-resistant isolates were also resistant to benzimidazole. These results show that failure of strobilurin fungicides to prevent leaf blight was caused by occurrence of strobilurin-resistant isolates. This appears to be the first report of strobilurin-resistant isolates of Pseudocercospora vitis.

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bendazim WP, Tebuconazole SC, Tebuconazole WG and Trifloxystrobin + Tebuconazole SC) out of them, showed effective suppression with the mycelium growth of pathogenic fungus, and were selected to test in vivo of the field condition. Six fungicides, Azoxystrobin SC, Dithianon WG, Polyoxin D zinc salt + carbendazim WP, Tebuconazole SC, Tebuconazole WG and Trifloxystrobin + Tebuconazole SC were significantly effective to protect anthracnose of Lycium chinese, the variety “Myeong-An”.

D-6

Timing and Sequence of Postharvest Fungicide and Biocontrol Agent Applications for Control of Pear Skin Sooty Disease. J.S. Lee , J.H. Song , Y.S. Cho , J.H. Choi , D.H. Kim and H.H. Park . Horticultural & Herbal Crop Environment Div. National Institute of 1

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Horticultural & Herbal Science, RDA, Suwon 440-706, Korea; Pear Experimental Station, National Institute of Horticultural & Herbal Science, RDA, Naju 523-820, Korea 2

Postharvest skin sooty dapple disease of pear fruit often originates at small stain symptoms that occur during harvest and handling. Experiments were conducted to characterize the effect of timing of application of skin sooty disease control materials, and to evaluate sequential postharvest applications of fungicides or fungicides and biocontrol agents. Fungicides and biocontrol agents were increasingly less effective when the period between harvest and application was prolonged. Thiabendazole (TBZ) applied to fruit without artificial wounding or inoculation effectively reduced skin sooty disease when applied within 3 weeks or 6 weeks in 2 years of study. TBZ, polyoxin B, and pyrimethanil were effective in controlling skin sooty disease at artificial wounds inoculated with Cladosporium tenuissimumup to 14 days after inoculation. Application of TBZ at harvest followed 3 weeks later by application of polyoxin B was superior to application of TBZ at harvest alone. One bacterial biocontrol agents reduced skin sooty disease at pear wounds inoculated with C. tenuissimum up to 14 days after inoculation with C. tenuissimum, but were ineffective when applied at 28 days after inoculation. Of possible sequential arrangements of fungicide and biocontrol treatments, application of the most effective material promptly after harvest generally resulted in the highest level of skin sooty disease control.

D-7

Selection of Bactericides for controlling Soybean bacterial pustule. Sung-Jun Hong , Yong-Ki Kim , Hyeong-Jin Jee , Bong-Choon 1

1

1

Lee , Young-Nam Yoon and Sung-Tae Park . Organic Agriculture 2

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Division, NAAS, RDA, Suwon 441-707, Korea; Department of Functional Crop Science, NICS, RDA, Miryang 627-803, Korea 2

D-5

Select of fungicides to control of Lycium Chinense Anthracnose caused by Colletotrichum spp. Han-Mo Koo , Seok-Jung Lee , 1

1

Yong-Jin Park , Chang-Ho Kim and Tea-Seok Oh . Department of 1

1

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1

Plant Resourses, Kongju National University, Yesan 340-702, Korea

Anthracnose caused by Colletotrichum spp. has been known as a significant disease which commonly infects to the fruits of Chinese matrimony(Lycium chinese) in the field conditions. Sellect effective fungicides for the control of Lycium chinese anthracnose, Six fungicides (Azoxystrobin SC, Dithianon WG, Polyoxin D zinc salt + car-

Bacterial pustule of soybean [Glycines max (L.) Merr.] caused by Xanthomonas axonopodis pv. glycines is one of the most prevalent bacterial diseases of soybean. Besides, this bacterium showed strong pathogenicity to the plants and distributed throughout Korea. However, no control measures including bactericides and resistant cultivars are available to control the disease in Korea. Therefore, this study is conducted to develop chemical control method for soybean bacterial pustule. The present study was undertaken to find out the growth inhibitory effect against bacterial pustule pathogen using bactericides (8 antibiotics, 2 copper compounds, quinoline, 18 agro-

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chemicals). Results of the antibiotics test showed that tetracycline and streptomycin sulfate significantly suppressed the growth of bacterial pustule pathogen. Also, application of oxolinic acid was found to be effective for pathogen inhibition. However, vancomycin, polymyxin B sulfate and copper compounds did not show the positive suppressive effect on growth of the pathogen. Among the eighteen agrochemicals, streptomycin sulfate + oxytetracyclin (18.8% + 1.5%) WP, oxytetracyclin (17%) WP and oxolinic acid (20%) WP were found to be effective for the inhibition of the pathogen in vitro. The selected agro-chemicals were also applied on soybean in field and their control effects of the soybean bacterial pustule were tested. The application of sreptomycin sulfate + oytetracyclin (18.8% + 1.5%) WP and oxytetracyclin (17%) WP to the natural infected soybean (Taekwangkong) showed high control value (above 70%). Therefore, it is concluded that bactericides used in this study showed strong inhibitory effect to soybean bacterial pustule and they can be recommend to farmers to control the bacterial pustule of soybean.

D-8

Control Effect of NaDCC for Major Diseases on Growing Stage in Pear. Deok-Hoon Yoon , Mi-Kyong Han and Ki-Woong Nam . 1

2

1,2

Research Institute for the Far East Asian Bio-Resources, Hankyong National University, Anseong 456-749, Korea; Department of Horticulture, Hankyong National University, Anseong 456-749, Korea 1

2

NaDCC(Sodium Di-Chloroiso Cyannurate) has been used as a food additive to inhibit a fruit rot during storage. A Pear Scab(Venturia nashicola) and a Pear Rust(Gymnosporangium haraeanum) are major diseases on growing stage in Pear, and, a chemical pesticide for these diseases are sprinkled more than 10 time per year in the fields. This study was conducted to check of chemical pesticide and to seek after prevention of the disease in environmental friendly. A NaDCC-1,000, NaDCC-1500, NaDCC-2000 diluted solution and difenoconazole4,000 diluted solution and no treatment were treated on growing stage in Pear. These were sprinkled 5 times in 10 days interval from the incipient stage of a disease. As a result, an incidence of the Pear Scab with no treatment was 3.5% in average against 1.5% in difenoconazole-4,000, 0.3% in NaDCC-1,000, 0.5% in NaDCC-1,500 and 0.7% in NaDCC-2,000. In case of an incidence of Pear Rust, no treatment showed 15.2% in average as compared with 8.7% in difenoconazole4,000, 7.5% in NaDCC-1,000, 7.6% in NaDCC-1,500 and 9.1% in NaDCC-2,000. A NaDCC-1,000 diluted solution, therefore, was showed a possibility to control both the Pear Scab and the Pear Rust. The damage from the NaDCC-1,000 diluted solution was not detected on Pear tree in this study.

D-9

Effect of NaDCC on Pollination and Germination on Flowering Stage in ‘Niitaka’ Pear. Deok-Hoon Yoon , Mi-Kyong Han , Hae1

2

Jun Park and Ki-Woong Nam . Research Institute for the Far East 2

1,2

1

Asian Bio-Resources, Hankyong National University, Anseong 456749, Korea; Department of Horticulture, Hankyong National University, Anseong 456-749, Korea 2

Pear tree is classified in a cross-pollination plant. However, an artificial-pollination has been widely operated due to a shortage of pollinators caused the worse of an environment. On the flowering stage of

Pear, moreover, a chemical spraying to prevent against disease and pest is limited to a chemical damage. To solve this problems, a NaDCC(Sodium Di-Chloroiso Cyannurate), as a food additive permitted by Korea FDA, has been used in this study. The NaDCC 1,000, 1,500 and 2,000 diluted solutions were sprinkled on the flower of ‘Niitaka’ Pear in the early stage of flowering and in the stage of full bloom. Then, a chemical damage and an incidence of pollination were investigated; besides, an incidence of germination of pollen with NaDCC spraying was investigated for Wonhwang, Chuhwang, Hwasan, Gamcheon, and Pungsu as pollinizers for ‘Niitaka’ Pear. An artificial pollination for Niitaka Pear was done on 18 April. In the early stage of flowering of Niitake Pear, an incidence of pollination was 98.7%, 98.7% and 99.3% with NaDCC 1,000, 1,500 and 2,000 respectively compared with 95.3% in no treatment. However, it was similar with no treatment in the stage of full bloom; 96.0%, 92.7% and 95.3% with NaDCC 1,000, 1,500 and 2,000 respectively. As a results of an incidence of germination of pollen for pollinizers, the Wonhwang germinated 70.4% with NaDCC 1,500 compared with 69.2% in no treatment, and the Gamcheon germinated 82.9% with NaDCC 2,000 compared with 64.9% in no treatment. Germination rates of Chuhwang, Hwasan and Pungsu with NaDCC were lower than no treatment in 60.9%-80.1%, 62.7%-68.9% and 35.8%-48.6% respectively. A chemical damage was not detected in all treatment in this study.

D-10

Silage Corn of disease and pest occurrence present conditions which it follows in fertilization of the phosphoric acid. Tea-Seok

Oh , Chang-Ho Kim , Han-Mo Koo , Seok-Jung Lee . Department 1

1

1

1

1

of Plant Resourses, Kongju National University, Yesan 340-702, Korea

The corn with the crops fertilizer plentifully in necessity the productivity of Amount of fertilizer qualitative seeds of the nitrogen and the phosphoric acid improve. It used Guano from the study which it sees and after increasing the commitment quantity of the phosphoric acid, disease and pest where it occurs productivity improvement of the corn and it investigated. The method seeding silage corns (Variety. Cheng An Ok) width (70cm) * distance (20cm), After setting encourage amount of fertilizer N-P-K-composts = 21-17-17-3,000kg of the corn (10a standards) with control, amount of fertilizer of the phosphoric acid after increasing at 20% unit, it investigated disease and pest where it occurs in the corn. During rearing term of the corn in southern corn leaf blight and Ostrinia furnacalis damage observe, It compared of Control that 40% southern corn leaf blight 21% and Ostrinia furnacalis above 27%, it was confirmed input of the phosphoric acid from treatment where it increases that the occurrence ratio increases.

D-11

Control efficacy of a new fungicide fludioxonil on lettuce gray mold according to several conditions. Kyoung Soo Jang, Song-Yi Baik, Jin-Cheol Kim, Young Ho Choi and Gyung Ja Choi. Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea

Fludioxonil is derived from the antifungal compound pyrrolnitrin produced by Pseudomonas pyrocinia and classified as a reduced risk fun-


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gicide by the US EPA. The efficacy of fludioxonil for the control of lettuce gray mold caused by Botrytis cinerea was evaluated under several conditions such as growth stages of host, inoculum concentrations, and amounts of potato dextrose broth (PDB) included in spore suspension of B. cinerea. At 4-leaf stage of lettuce plants, fludioxonil applied at 2 g/ml was more effective for the control of gray mold than at 5- and 6-leaf stages. However, fludioxonil at more than 10 g/ml provided similar control activity in all growth stages of lettuce tested. The fungicide (10 and 50 g/ml) also gave excellent control of gray mold on lettuce seedlings inoculated with spore suspensions of B. cinerea (2.5×10 to 2×10 spores/ml). But, control efficacy of fludioxonil (2 g/ml) was negatively correlated with concentrations of inoculum. Addition of PDB in spore suspension of B. cinerea resulted in higher disease severity than non-treated control. By inoculating spore suspension including 0.5% PDB, the fungicide gave the most control activity on the disease, followed by 1% and 2% PDB. The results suggest that fludioxonil has potential to control gray mold of lettuce, but the fungicide at a concentration having moderate activity may represent low control efficacy on the disease under some conditions.

Bacterial canker and wilt, caused by Clavibacter michiganensis subsp. michiganensis and Ralstonia solanacearum, are severe disease in tomato. To develop a natural fungicide, methanol extracts of 333 plants were screened for in vitro anti-bacterial activity against Clavibacter michiganensis subsp. michiganensis and Ralstonia solanacearum. Among them, three plant extracts showed high diseasecontrol efficacy of more than 90% against at least one of six plant diseases. The methanol extracts of Ruta grabeolens L., Polygala tenuifolia Willd. and Galium verum var. asiaticum displayed the highest anti-bacterial activity against baterial canker. Also, the methanol extracts of Isatis tinctoria L. and Rumex acetosa L. displayed the highest anti-bacterial activity against baterial wilt. Further studies on the characterization of anti-bacterial subtances in plant extracts are underway and their disease-control effects should be performed under greenhouse and field conditions.

D-12

D-14

5

6

Control efficacy of systemic treatment against clubroot in Chinese cabbage field. Jae Goon Lee, Moo Ill Yeo, Ho Cheol Shin and In Cheon Hwang . Central Research Institute Kyung Nong Corpora1

1

tion, Gyeongju 780-110, Korea

Clubroot caused by Plasmodiophora brassicae is one of the most destructive disease in Chinese cabbage field. Until now, Chemical control of clubroot was soil incorporation treatment before transplanting by Flusulfamide DP/GR, Fluazinam DP or dipping treatment before transplanting by Cyazofamid SC. As Plasmodiophora brassicae lived in soil, traditional control efficacy was affected by soil condition and incorporation depth in soil. In this study, we investigated control efficacy of systemic treatment (soil incorporation treatment+dipping treatment) against clubroot in order to establish more stable control method than traditional methods. Fungicide used in field trial were Flusulfamide+Flutolanil 0.3+1% DP and Cyazofamide 10% SC. Field trial was carried out in Gyeongju, Korea. Efficacies were evaluated by disease degree and disease rate. Disease degree and disease rate of untreated plot were 39.9% and 65.6%. Traditional treatment by Cyazofamide SC, Flusulfamide+Flutolanil DP and Flusulfamide DP showed 11.9%, 19.5%, 20.1% of disease degree and 11.1%, 31.0%, 23.7% of disease rate, respectively. Systemic treatment (soil incorporation treatment+dipping treatment) showed 11.4% of disease degree and 9.8% of disease rate. These results indicated that efficacy of Systemic treatment (soil incorporation treatment + dipping treatment) was higher than traditional treatment against clubroot. In addition, half application of Cyazofamid SC in systemic treatment showed 15.4% of disease degree and 9.8% of disease rate. Our results suggested that systemic treatment (soil incorporation treatment + dipping treatment) with Flusulfamide+Flutolanil DP and Cyazofamide SC might be more effective control method against clubroot.

Han, Dae-Hyun Kim, Jung-sup Lee and Jeong-Hwan Hwang. Horti-

cultural & Herbal Crop Environment Division, National Institute of Horticultural & Herbal Science, RDA, Suwon 441-440, Korea

Effects of A Orysastrobin Pre-mix Formulation with Thiamethoxam(COMBINE ) on Several Rice Diseases according to the Nusery-box Treatment. Dae-Yong Um, Lae-Hong Cho, Hee®

Jun Kwon, Young Kwon Kim and Tae-Joon Kim. Crop Protection Research Team AgroLife Research Institute, Dongbu HiTek Co. Ltd., Moonjidong, Daeduck Science Town, Daejeon, 305-708, Korea

The control released pre-mix formulation of 3.5% orysastrobin with 1.0% thiamethoxam has been commercialized for controlling rice diseases through nursery box treatment. Application of this pre-mix granule formulation showed around 80% control efficacy against rice leaf blast, neck of panicle blast, and rice sheath blight with 50 g/ box(30×60 cm ), which consists of orysatrobin and thiamethoxam at 350 g ai/ha and 100 g ai/ha, respectively. With the same application rate, small brown planthopper known as an important vector of rice stripe virus (RSV) was perfectly controlled. This result suggested that the population of rice stripe disease caused by RSV could be effectively reduced in a paddy rice cultivation. Simultaneously, this premixture didn't show any phytotoxicity on rice plants in nursery boxes through various application methods such as hand scattering and soil incorporating at the time of the rice box preparation. In addition, the rice plant was very safe through different application time of 7 days and the day before transplanting. We recommend that the controlreleased granule type of 3.5% orysastrobin with 1.0% thiamethoxam is a useful tool controlling several rice diseases for the farmers. Moreover, it could be both effective and labor-saving method when using the spreading machine for controlling the diseases in the rice cultivation. 2

D-15

Chemical control efficiency of Alternaria leaf blight diseased on Korea carrot. Kyeonghun Park , Kyusik Jeong , Eunhee Soh , 1

D-13

Anti-bacterial effects of Plant extracts Against Bacterial canker and wilt of Tomato. You-Kyoung Han, Jong-Han Park, Kyung-Sook

2

2

Byungseok Kim , Seokcheol Kim and Dooho Kim . National Acad1

1

1

1

emy of Agricultural Science, RDA, Suwon 441-707, Korea; Variety Testing Division, Korea Seed & Variety Service, Suwon 443-400, Korea 2

291


Leaf blight of carrots caused by Alternaria dauci(Kuhn) Groves and Skolko is often the most serious disease affecting carrots in the southern United States and causes serious losses in midwestern and eastern states and in other areas of the world. Incidence and severity of Alternaria leaf blight on Korea carrot crops increased recently. Fungicide application to control disease development has become more common, but there is no information on the effects of A. dauci infection on the yield and quality of carrot. This study will give useful information about time and frequency of fungicide applications for more effective chemical control of Alternaria leaf blight. The saccharinity was closely related with progress of disease in Korea carrot. In higher saccharinity carrot, progress of disease was reduced and in lower saccharinity carrot, it was increased. The proper time of fungicide applications for the chemical control of Alternaria leaf blight would be early in July in Jinbu and early in august in Daegwallyeng, respectively and the treatment frequency of fungicides showed the highest control efficiency to treat 7 times at 7 days intervals. Propineb, difenoconazole and copper hydroxide provided 86.9%, 85%, 77.6% in control efficiency, respectively, in the field experiment with carrot 'mussang- 5chon'. The disease control with fungicides was increased production yield and the efficiency of chemical control to reduce Alternaria leaf blight disease was high.

D-16

Screening of Seed Coating Pesticide for Control of Damping-off caused by Rhizoctonia solani on Panax ginseng. Jung-Sup Shin *, 1

Seung-Hwan Lee , Hye-Sun Cho , Dae-Hui Cho and Hee-Young Jung . Ginseng Research, KT&G Central Research Institute, Dae1

2

1

Lee, Yong Hwan Lee, Inn-Shik Myung, Dong-Soo Ra and Se-Weon Lee. Agricultural Microbiology Division, National Academy of Agricultural Science, RDA, Suwon 441-707, Korea

Recently, incidence of rice bakanae disease caused by the ascomycete fungus Fusarium moniliforme has increased rapidly in Korea. Prochloraz and hexaconazole are the popular control agents for suppression of the fungus. Prochloraz and hexaconazole inhibit CYP51 gene product, the cytochrome P450 lanosterol 14α-demethylase, which is the essential part of erogosterol biosynthetic pathway. To evaluate the resistance to prochloraz and hexaconazole, EC values were investigated using the agar dilution method. EC value of F. moniliforme to prochloraz ranged from 0.002 to 1.548 µg/mL with an average of 0.17 µg/mL. EC values to hexaconazole ranged 0.010 17.253 µg/mL, and the average was 2.43 µg/mL. The resistant baseline of F. moniliforme to prochloraz and hexaconazole was 0.3 µg/mL and 5.0 µg/mL, respectively. Based on the resistant baseline, the proportion of resistant isolates to prochloraz and hexaconazole were 12.9 % and 10.1 %, respectively. Hexaconazole resistant isolates were distributed nationwide in Korea and prochloraz resistant isolates were also detected nationwide except Chungbuk and Jeonbuk provinces. Jeonnam province showed the highest proportion of resistant isolates to both prochloraz (66.7%) and hexaconazole (25%), however there was no evidence of cross-resistance between prochloraz and hexaconazole. Comparative analysis of CYP51 is in progress to validate the genetic difference between the resistant and the susceptible pathogens. 50

50

50

1

1

jeon 305-805, Korea; School of Applied Biosciences, Kyungpook National University, Daegu 702-701, Korea

D-18

Damping-off caused by Rhizoctonia solani is an important disease causing considerable damage on ginseng of nursery bed. Tolclofosmethyl was used against this disease for 20 years in general but 6 years after coating seeds with tolclofos-methyl were planted, this chemical was still remained in ginseng root. So we need to find alternative pesticide. This study is about the activities of 4 fungicides (Fludioxonil, Flutolanil, Pencycuron, Thifluzamide) which were registered as drenching type fungicide on R. solani. Mycelial growth inhibition rates of each fungicide were tested in vitro method for testing soil fungicides (Zentmyer, 1955). Fludioxonial showed 100% inhibition activity, Thifluzamide, Flutolanil and Pencycuron showed 85%, 98% and 75%, respectively. Experimental plots (180×90cm) of ginseng nusery bed were artificially infested with 50g fresh weight of inoculum. Ginseng seeds wer wated with the fungicides and seeded to the field. Fludioxonil, flutolani, pencicuron, and thifluzamide reduced the incidence of damping-off by 71%, 57%, 54% and 29%, respectively. The residue of Fludioxonil which showed the highest mycelial growth inhibition activity in vitro and in vivo was from 0.44 to 1.15 mg/kg after 11months cultivation, it comes to the conclusion that the remaining residue will be adapt to regional MRLs after 6-year cultivation and fludioxonil is able to be used instead of Tolclofos-methyl against damping-off.

Park, Yong Hwan Lee, In-Sik Myeong, Dong-Su Ra and Seong-Suk Han. Agricultural Microbiology Division, National Academy of Agri-

2

Effects of combined solutions of several fungicides on mycelial growth and spore germination of Fusarium fujikuroi. Woo-Sik cultural Science, RDA, Suwon 441-707, Korea

We investigated the effects of various concentrations of two fungicides combined solution on spore germination and mycelial growth of resistant and sensitive strains of Fusarium fujikuroi to prochloraz. Mycelial growth of sensitive strain was inhibited 100% at 10 µg/ml of prochloraz and 65.8% at 10 µg/ml of hexaconazole. But, that of resistant strain was inhibited 95.6% at 30 µg/ml of prochloraz and 55.8% at 10 µg/ml of hexaconazole. Interestingly, mixed solution of 10 µg/ ml and 0.1 µg/ml of prochloraz and hexaconazole inhibited 100% of mycelial growth of sensitive strain and mixed solution of 10 µg/ml and 1.0 µg/ml of prochloraz and hexaconazole inhibited 100% of that of resistant strain. While fludioxonil showed 99.6% of inhibition rate at 80 µg/ml on mycelial growth of resistant strain, mixed solution of 20 µg/ml and 5 µg/ml of prochloraz and fludioxonil inhibited 100% of that of resistant strain.

-1

D-19

Sensitivity of boscalid-resistant and -sensitive isolates of Corynespora cassiicola and Podosphaera xanthii to a novel succinate dehydrogenase inhibitor fluopyram. Hideo Ishii , Takuya 1,2

D-17

Resistance of Fusarium moniliforme Causing Rice Bakanae Disease to Demethylation Inhibitor Fungicides. Jae-Yun Jeong, Zu-An

Miyamoto , Shingo Ushio and Makoto Kakishima . University of Tsukuba, Tennodai 1-1-1, Tsukuba, Ibaraki 305-8571, Japan; National Institute for Agro-Environmental Sciences, Kannondai 3-13

2

4

1

1

Abstract of Presentations 3, Tsukuba, Ibaraki 305-8604, Japan; Horticultural Institute, Ibaraki Agricultural Center, Kasama, Ibaraki 319-0292, Japan; Chiba Prefectural Agriculture and Forestry Research Center, Midori, Chiba 266-0006, Japan 3

4

Recently in Japan, isolates resistant to boscalid, a succinate dehydrogenase inhibitor (mitochondrial complex II inhibitor, SDHI), have been detected in Corynespora cassiicola and Podosphaera xanthii, the pathogen of Corynespora leaf spot and powdery mildew disease on cucumber, respectively. Resistant isolates of C. cassiicola represent a serious problem in disease control and are widely distributed at present. In this study, the sensitivity of these fungal pathogens to fluopyram, a novel SDHI under development, was tested by bioassays. Boscalid-resistant isolates of C. cassiicola were clearly distinguished from sensitive isolates when comparing their mycelial growth on YBA agar medium supplemented with this fungicide. In contrast, the difference of fluopyram sensitivity between boscalid-resistant and sensitive isolates was much less if any. Interestingly, the growth of both boscalid-resistant and -sensitive isolates was suppressed on fluopyram-amended YBA agar medium. In addition, isolates showing very high resistance to boscalid in a mycelial growth test on a culture medium were controlled well by fluopyram in an inoculation test using potted cucumber plants. Sensitivity of P. xanthii isolates to fluopyram was further examined using cucumber leaf disc assays and pot experiments. Although boscalid hardly controlled powdery mildew development when very highly boscalid-resistant isolates were employed for inoculation, fluopyram still exhibited high control efficacy against boscalid-resistant as well as -sensitive isolates of this fungus. Differential sensitivity to boscalid and fluopyram found in these two pathogens may indicate the involvement of a distinct mode of action for fluopyram compared to boscalid. This finding may lead to the discovery of unique SDHIs in the future.

292

was higher for technique II than the other methods; this technique detected concentrations as low as 10 conidia/ml, similar to that for the conventional assay. The results suggest that the technique II assay substantially enhanced sensitivity of real-time PCR for detecting B. cinerea on symptomless flowers.

E-2

Development of PCR method for detection of Pectobacterium carotovorum. Jeong-Gu Kim , Ji-Na No , Mi-Sun Yoo , Dong-Suk 1

2

2

Park and Byoung-Su Yoon . Genomics Division, National Academy of Agricultural Science, Rural Development Administration, Suwon, 441-707, Korea; Department of Life Science, College of Natural Science, Kyonggi University, Suwon, 443-760, Korea; National Agrobiodiversity Center, National Academy of Agricultural Science, Rural Development Administration, Suwon, 441-707, Korea 3

2

1

2

3

New PCR method was developed to detect for Pectobacterium carotovorum subsp. carotovorum which is the causative agent of soft rot in Brassica pekinensis. A specific detection primer based on Lytic murein transglycolase gene was designed and evaluated. Using ERB3_F(5'-TGC GAC ACC TCC TCA TCA CG-3') and ERB3_R(5'-CTT ATC ACG CTG TAA CCA GC-3') primers, 437 nucleotides long fragment was specifically amplified. In result of specific PCR to 63 kinds of different organisms, PCC specific amplification was revealed 52 strain except KACC10435, KACC13958, KACC13968. Total 55 strains belong to Pectobacterium carotovorum or Pectobacterium carotovorum subsp. carotovorum. The other hand, 8 kinds of organisms including Chinese cabbage and potato did not detected. Optimal condition of ERB 3 F/R primer was established at 58 C in annealing temperature and 15mM MgCl concentration. In the PCR assays with serially diluted template, the specific PCR showed high sensitivities which were able to detect 10 copies. Also, this method can detect not only to be used for DNA condition but also to be applied for real field samples. This PCR method is expected to be useful for specific detection of Pectobacterium carotovorum. o

2

3

E-1

Development of a real-time PCR assay for detection of Botrytis cinerea in strawberry flowers. Myeong-Hyeon Nam , Yun-Gyu 1

Nam , Jung-Young Song , Mark Gleason and Hong-Gi Kim . 1

2

3

2

Nonsan Strawberry Experiment Station, Chungnam ARES, Nonsan, 320-862, Korea; Department of Applied Biology, Chungnam National University, Daejeon 305-764, Korea; Department of Plant Pathology, Iowa State University, Ames, IA 50011, USA 1

2

3

E-3

Complete nucleotide sequence of Japanese strains of Barley yellow mosaic virus and discrimination by multiplex RT-PCR.

Hisashi Nishigawa , Mari Yumoto , Tomomi Hagiwara , Toshinori Sotome , Tsuneo Kato , Koji Kimura and Tomohide Natsuaki . 1

3

Detection of Botrytis cinerea on symptomless strawberry flowers is critical to improving management of anthracnose fruit rot. A realtime PCR assay with BcTaq424f/BcTaq491r/BcFamP (Cadle-Davison, 2008) was used for detecting the pathogen in vivo. These probes were specific for B. cinerea from strawberry; detection thresholds were 100 fg of genomic DNA from mycelium and 10 conidia/ml and conidial suspensions, respectively. Sensitivity was assessed for three ways of handling inoculated tissue. Strawberry flowers were sprayinoculated with 1.0 to 10 conidia/ml. Petal, stamen and sepal tissues were sampled and placed in eppendorf tubes. To dislodge the pathogen, DNA was either extracted directly (technique I), or prior to DNA extraction, tissues were frozen for 12 h, incubated for 2 days at 27°C with 50 ul of sterile distilled water, and then sonicated and agitated (technique II), or frozen for 12 h, incubated for 2 days at 27°C and then sonicated and agitated (technique III). DNA was extracted using CTAB+1% polyvinylpyrrolidone. Detection sensitivity for all tissues 3

5

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4

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5

1,2

Center for Bioscience Research & Education, Utsunomiya University, Tochigi 321-8505, Japan; Faculty of Agriculture, Utsunomiya University, Tochigi 321-8505, Japan; Tochigi Prefectural Agricultural Experiment Station, Tochigi 320-0002, Japan; Tochigi Prefectural Agricultural Management Division, Tochigi 320-8501, Japan; Agricultural Promotion Division, Yamaguchi Prefectural Government, Yamaguchi 753-8501, Japan 1

2

3

4

5

Complete nucleotide sequences of four strains (I, III, IV and V) of Barley yellow mosaic virus (BaYMV) isolated in Japan were determined. The length of the genome was the same among these strains; RNA1 was 7,642 nt and RNA2 was 3,585 nt. The molecular phylogenetic analysis showed that strains II to V and the Korean strain (AF536958 and AF536957) formed one cluster, and strain I, Chinese and European strains formed the other cluster. Amino acid sequences of each viral gene product were compared among strains. The

293


sequences of the VPg protein showed less identity among almost strains (less than 92%) than those of other proteins (more than 93%). VPg is thought to be involved in interactions with host factors, especially initiation factor 4E (eIF4E) or eIF(iso)4E, and infection. Concerning Japanese BaYMV strains, even though only VPg does not always determine phenotypic symptoms to barley strains, the fact that there were more mutations in VPg than other genes prompted us to establish a multiplex reverse transcription polymerase chain reaction (RT-PCR) method to discriminate among Japanese BaYMV strains. We designed strain-specific primers from the VPg sequences and targeted single nucleotide polymorphisms (SNPs). The PCR results for strain identification corresponded with pathogenicity of each strain.

E-4

Developing Molecular Markers for the Identification of Blue Stain Fungi, the Ophiostoma piceae Complex. Min Woo Hyun , 1

Dong Yeon Suh , Yeo Hong Yun and Seong Hwan Kim . 1

1

1,2

Department of Microbiology and Institute of Basic Sciences, Dankook University, Chungnam, 330-714. Korea

1

2

The species of Ophiostoma piceae complex include economically important species known as serious tree pathogens and agents of causing blue stain in lumber and timbers. Particularly, in the complex group, O. piceae, O. quercus, O. floccosum and O. setosum are represented agents that commonly occur. Since these species are morphologically so similar, it is hard to distinguish each species using traditional methods. To develop more easy and fast method for their differentiation, we designed molecular marker based on-tubulin gene sequences. The species-specific primers identified O. piceae (45 isolates), O. quercus (9 isolates), and O. floccosum (24 isolates), respectively, from 107 isolates of unidentified fungi that were isolated from blue stained wood in Korea. In addition, the designed primers did not react with other Ophiostoma species (O. ips, O. minus, O. cannum) and Leptographium spp. and other wood inhabiting fungi including Penicillum spp. and Trichoderma spp. Therefore, it is expected that the species-specific primers are useful for the rapid detection and classification of the four blue-stainers of O. piceae complex.

E-5

Application of automatic multiplex real-time RT-PCR method for the fast quarantine testing of 20 plant pathogenic viruses, especially for the Cucurbitaceae family. Jin Hyuck Kim , Hyun Hee 1

Kim , Ki Hyun Ryu , Ji Yeon Kwon , Seon Mi Choi , Sung Won Hong , Soo Bok Kim , Myo Ah Baek and Yong Suk Nam . 2

3

1

3

1

1

1

1

KOGENEBIOTECH CO., LTD., 1203 Woolim Lions Valley C-dong, 371-28 Gasan-dong, Geumcheon-gu, Seoul, Korea; Department of Life Science, Sahmyook University, Seoul 130-650, Korea; Department of Horticulture, Seoul Women's University, Seoul 139774, Korea 1

2

CGMMV, CMMoV, HLSV, KGMMV, MarMV, ORSV, PMMoV, RMV, SFBV, TMGMV, TMV, ToMV, ZGMMV, WMV, ZYMV, MNSV, and PRSV - responsible for infection of the ornamental plants and the plant family Cucurbitaceae such as Citrullus vulgaris, Cucumis sativus, Cucumis melo var. makuwa, and so on. The assay targets the each virus-specific region compared with full genomes of viruses. To allow multiplex detection of 20-pathogens, specific primers and TaqMan probes labeled with the various fluorescent markers, such as FAM, VIC, NED, and Cy5, were designed. Internal positive control targeted to plant, designed as 26s rRNA region of chloroplast, was codeveloped for 20 pathogenic plus/minus assay and single and multiplex assay were performed and optimized on AB7500. Furthermore, Automatic Real-time PCR Analysis Software was developed which can analyze data extracted after real-time PCR run. Ultimately the assay was constructed as quadruplex system including three viruses and IPC can be simultaneously detected on one tube, the same as seven reactions for twenty viruses in one multiplex set. Automatic analysis software is an integral part of the developed detection system and can decide which virus is contaminated within 5 minutes. Overall, the developed system is reliable and easy to use, and the entire procedure can be completed in less than 2 hours. Automatic multiplex real-time RT-PCR system can simplify plant quarantine testing and be efficiently applied in case of multiple infection.

E-6

Developing Molecular Markers for the Identification of Blue Stain Fungi, the Ophiostoma piceae Complex. Min Woo Hyun , 1

Dong Yeon Suh , Yeo Hong Yun and Seong Hwan Kim . 1

1

1,2

Department of Microbiology and Institute of Basic Sciences, Dankook University, Chungnam, 330-714, Korea 1

2

The species of Ophiostoma piceae complex include economically important species known as serious tree pathogens and agents of causing blue stain in lumber and timbers. Particularly, in the complex group, O. piceae, O. quercus, O. floccosum and O. setosum are represented agents that commonly occur. Since these species are morphologically so similar, it is hard to distinguish each species using traditional methods. To develop more easy and fast method for their differentiation, we designed molecular marker based on -tubulin gene sequences. The species-specific primers identified O. piceae (45 isolates), O. quercus (9 isolates), and O. floccosum (24 isolates), respectively, from 107 isolates of unidentified fungi that were isolated from blue stained wood in Korea. In addition, the designed primers did not react with other Ophiostoma species (O. ips, O. minus, O. cannum) and Leptographium spp. and other wood inhabiting fungi including Penicillum spp. and Trichoderma spp. Therefore, it is expected that the species-specific primers are useful for the rapid detection and classification of the four blue-stainers of O. piceae complex.

3

It is always significant subject to develop more rapid and easy method simultaneously detecting much more pathogenic virus without false positive results because it offers the opportunity to increase the efficiency of plant quarantine testing. In this context, an automatic multiplex real-time PCR system has been developed for the specific detection of representative 20-plant viruses - BPeMV, CFMMV,

E-7

Multiplex RT-PCR detection of Cucumber mosaic virus, Ribgrass mosaic virus and Turnip mosaic virus in Chinese cabbage (Brassica campestis ssp. pekinensis). Hye Jeong Kim , Chul Young Park , 1

1

Jin Sung Hong , Jeong-Soo Kim , Ki Hyeon Ryu , Jang Kyung Choi and Sang Yong Lee . Division of Forest Resources Protection, Kang3

4

1

3

2

1

won National University, Chuncheon 200-701, Korea; Division of Bio-Resources, Kangwon National University, Chuncheon 200-701, 2

Abstract of Presentations Korea; Division of Environmental and Life Sciences, Seoul Women’s University, Seoul 139-744, Korea; National Institute of Agricultural Science & Technology, Rural Development Administration, Suwon 441-707, Korea 3

4

E-9

Southern Hybridization Detection of Rhizina undulata in Soil Using DIG-labelled cDNA Probes Synthesized by ITS - specific primers. Sun Keun Lee , Hwee Lee , Jong Kyu Lee , Seung Kyu 1

o

o

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E-8

Molecular Detection of Phytophthora capsici in Soil and Pepper Plants by Real-Time Polymerase Chain Reaction Using SYBR Green. Jeong Young Song *, Jin Ha Lim , Myeong Hyeon Nam , 1

1

2

You Ri Choi and Hong Gi Kim . Dept. of Agricultural Biology, 1

1

1

Chungnam National University; Nonsan Strawberry Experiment Station, Chungnam ARES 2

Phytophthora blight of pepper, caused by Phytophthora capsici, is widespread and destructive to pepper (Capsicum annuum L.) in many areas of the world. A specific and sensitive real-time PCR assay using SYBR green was developed for the detection and identification of P. capsici. The species-specific primer pairs for P. capsici were designed from the intergenic spacer (IGS) and retrotransposable element region. PCR amplification with these primer sets resulted in amplification of products of approximately 100 bp exclusively from isolates of P. capsici. The sensitivity limit of the primer sets were 10-100 fg of genomic DNA of P. capsici, approximately and the real-time PCR assays differentiated P. capsici from the 30 species of Phytophthora and other fungi tested. The assays also confirmed the pathogen in infected plant materials and infested soil samples from pepper fields. P. capsici was isolated on PARPH media from all plant samples which were quickly detected using NaOH method for DNA isolation and real-time PCR assays. The PCR assays could be used as an accurate and sensitive method for detecting P. capsici in soil and pepper plants.

1

1

Lee , Kyung Hee Kim and Sang Yong Lee . Division of Forest 2

Rapid and reliable detection techniques are very important to the control of plant viral disease. Cucumber mosaic virus (CMV), Ribgrass mosaic virus (RMV) and Turnip mosaic virus (TuMV) are the major viruses of Chinese cabbage (Brassica campestris ssp. Pekinensis) in Korea. Multiplex reverse transcriptase polymerase chain reaction (RT-PCR) has become a popular technique since it provides a quick, reliable and low-cost method for simultaneous detection of multiple viruses. The multiplex RT-PCR assay was developed to detect three viruses infecting Chinese cabbage. Virus-specific primers (C2R4T8) were designed for coat protein genes of each of the three viruses. Each primer produced specific amplicons with a size of 238 bp (CMV), 433 bp (RMV) and 819 bp (TuMV), respectively. The optimal annealing temperature for each viral primer pair was determined by gradient multiplex PCR assays at 9 different annealing temperatures (50~54, 56, 58, 60 and 62 C) in the mixture of 3 viral-cDNA templates. The best amplification efficiency for triple targets was obtained at a annealing temperature of 56 C. The primer concentrations (10, 50 and 100 pM) in the mixture of 3 viral-cDNAs did not effected to the PCR amplicon of CMV. According to the increment of the primer concentration, the PCR amplicons of TuMV were decreased while those of RMV were increased. Dilution end point of the cDNA from RT reaction, which was possible to detect 3 viruses by PCR, was about 10 . The sensitivity of the multiplex RT-PCR was similar to the sensitivity of simplex RT-PCR to detect 3 viruses. This rapid and specific multiplex RT-PCR assay could be provided an effective and reliable molecular diagnostic tool for the detection of 3 viruses infecting in Chinese cabbage.

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2

1

1

Resources, Kangwon National University, Chuncheon 200-701, Korea; Korea Forest Research Institute, Seoul 130-712, Korea 2

Rhizina root rot (Rhizina undulata Fr ex Fr.) is a soil-borne disease occurring on coniferous trees, and causes severe damages on coastal pine forests in Korea. To control forest diseases effectively, rapid and efficient detections of the causal pathogen is the first and very important step. Southern hybridization is one of rapid and effective diagnosis tools to detect specific gene. Dot blotting based on southern hybridization was applied to detect R. undulata in Pinus thunbergii forest soil in Korea. DIG (Digoxigenin)-labeled cDNA probe specific to R. undulata was synthesized by ITS1 and ITS4, which are common sequence in the ITS region of true fungi. The PCR product of R. undulata amplified by ITS1 and ITS4 was about 585 base pair. Forest soil collected from healthy stand was mixed with cultured mycelium of R. undulata, and PCR products were amplified from 0.4g of the soil-mycelium mixture by ITS1 and ITS4 primers. The dot hybridization using the DIG-labelled cDNA probe was reacted specifically to the PCR products amplified from the soil-mycelium mixture of R. undulate isolates. The dot blot hybridization could detect as low as 100pg mycelium of R. undulata in 100g soil. The dot blot hybridization using the DIG-labelled cDNA probe by ITS primer pairs (ITS1 and ITS4) was also applied to the detection of R. undulata in soils collected from P. thunbergii forest infested with the disease at Gangneung, Gangwon Province. R. undulata was mainly detected by the dot blot hybridization in the soil samples of 0 ~ 10 cm depth where dead trees and fruit bodies of R. undulata were present. The dot blot hybridization protocol could be used successfully to detect R. undulata from forest soils.

E-10

Biochemical and Molecular Characteristics of a Causal Agent of Bacterial Spot of Tomato from Korea. Sun Young Moon, Inn-Shik

Myung*, In Ho Jeong, Young-Kee Lee, Young Hwan Lee, Se Weon Lee and Dong Soo Ra. Agricultural Microbiology Division, National Academy of Agricultural Science, Rural Development Administration, Suwon 441-707, Korea

Biochemical, molecular and pathological characteristics of a causal agent of bacterial spot of tomato from Korea were investigated. The isolates were gram-negative aerobic rods with a single flagellum. Colonies were yellow and raised with smooth margins on peptone sucrose agar. Starch and pectate hydrolysis were positive. Phenotypic tests were performed using API 20 Non Enterobacteriaceae Identification Kit. The physiological and biochemical characteristics of the strains were similar to those of the standard reference strain of Xanthomonas euvesicatoria (ICMP109). The isolates were positive for esculin hydrolysis, gelatin liquefaction, citrate utilization, acetoin production, malonate utilization, esculin hydrolysis, β-galactosidase activity, and assimilation of D-glucose, D-mannose, N-acetyl-glucosamine, D-maltose, malic acid, citrate. Negative reactions were oxidase, reduction of nitrates to nitrites, indole production, fermatation, arginine dihydrolase, urease production, and assimilation of LTM

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arabinose, D-mannitol, gluconate, capri acid, adipic acid, phenylacetic acid. However, the 16S rRNA gene (rrs) and the intergenic spacer located between rrs and the 23S rRNA gene, and the partial sequences of gyrB of the isolates shared 100% identity with the analogous sequences of X. perforans. The isolates were pathogenic to tomato (Solanum lycopersicun cv. Seokwang) and hot pepper (Capsicum annuum cv. Daekwang).

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Biochemical and Molecular Characteristics of a Causal Agent of Bacterial Stalk Rot of Sweet Corn from Korea. In Ho Jeong, Inn-

Shik Myung*, Sun Young Moon, Wan Gyu Kim, Se Weon Lee, Young Hwan Lee, Young-Kee Lee, Hong Shik Shim and Dong Soo Ra. Agricultural Microbiology Division, National Academy of Agricultural Science, Rural Development Administration, Suwon 441707, Korea

Biochemical and molecular characteristics of a causal agent of bacterial stalk rot of sweet corn from Korea were investigated. The bacterial isolates exhibited the biochemical characteristics of the family Enterobacteriaceae. They were Gram-negative, oxidase negative, catalase positive, fermentative, rod shaped, motile, facultatively anaerobic. All isolates were preliminary identified as Pectobacterium chrysanthemi with the Biolog Microbial Identification System, version 4.2 and with the PCR products acquired using primers by Nassar et al. Phenotypic tests were performed using Hi25 27 Enterobacteriaceae Identification Kit. The physiological and biochemical characteristics of all strains were similar to those of the reference strains of Dickeya chrysanthemi biovar chrysanthemi and D. chrysanthemi biovar. parthernii. All the isolated strains were positive for o-nitrophenyl-n-D-galactopyrano side, nitrate reduction, citrate utilization, acetoin production, malonate utilization, esculin hydrolysis, and utilization of melibiose, saccharose, glucose. Negative reactions were oxidase, lysine decarboxylase, ornithine decarboxylase, urease, deamination, H S production, indole production, and utilization of adonitol, rhamnose, trehalose, latose. Colonies were variable in methyl red and utilization of cellobiose, raffinose. The 16S rDNA region of the six isolates was sequenced using universal primers and compared with the region of reference strains of the Genus Dickeya. The isolates were clearly separated from the Genus Dickeya characterized previously, and into two groups in a phylogenetic tree generated with the neighbor- joining method in MEGA.

developed seven SCAR markers specific to P. katsurae generated by random amplified polymorphic DNA (RAPD), and assessed the potential of the SCAR markers as a tool for identifying P. katsurae. RAPD of total DNA extracted from P. katsurae was analyzed by random decamer primers (Operon, Inc.). RAPD of 16 Phytophthora species (P. citricola, P. cactorum, P. cinnamomi, P. cambivora, P.

lateralis, P. boehmeriae, P. citrophthora, P. cryptogea, P. drechsleri, P. nicotianae, P. gonapodyides, P. infestans, P. melonis, P. erythroseptica, P. sojae, P. palmivora) were also analyzed as the control. RAPD profiles were specific to each Phytophthora species, and seven DNA fragments distinguishing P. katsurae from other 16 Phytophtora species were screened. The P. katsurae-specific DNA fragments were

eluted from agarose gel using a QIAquick Gel Extraction Kit (QIAGEN Co.), ligated in a pGEM-T easy vector (Promega Co.), and cloned in competent Escherichia coli cells. Plasmid DNAs from transformed cell were purified by Wizard Miniprep DNA Purification System (Promega Co.). Purified plasmid DNAs were sequenced by ABI PRISM 3730XL Analyzer Sequencer (Applied Biosystems Co.). Seven primer pairs (OPC1F/OPC1R, OPC1-1F/OPC1-1R, OPC3F/OPC3R, OPC4F/OPC4R, OPC4F/OPC4-1R, OPD9F/ OPD9R and OPD10F/OPD10R) from the sequence analysis of the SCAR markers were designed. All of 7 primers were specific for P. katsurae in PCR detection and did not produce any P. katsurae-specific PCR amplicons from other 16 Phytophthora species used as the control. This study shows that SCAR markers must be a useful tool for rapid and effective detection of P. katsurae. TM

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Application of Sequence Characteristic Amplification Regions (SCARs) Markers for Detecting Phytophthora katsurae, the Fungal Pathogen of Chestnut Ink Disease in Korea. Hwee Lee, Sun keun Lee, Jong Kyu Lee and Sang Yong Lee. College of Forest and

Environmental Sciences, Kangwon National University, Chuncheon, 200-701, Korea Phytophthora katsurae is the fungal pathogen responsible for chest-

nut ink disease, which was first reported in Korea at 2005. Sequence characteristic amplification regions (SCARs) marker, one of effective and accurate tools for microbial identification, was applied for the rapid and accurate identification of P. katsurae. In this study, we

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Diagnosis of Xanthomoans oryzae pv. oryzae from Rice leaf Using Duplex primer. Tae Hwan Noh , Mi Hyung Kang , Du Ku Lee , 1

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Young Jin Park , Jong Chul Park , Su Joa Seo , Geon Hwi Lee , Si Ju Kim , Byoung Moo Lee and Hyung Moo Kim . National Institute of 2

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Crop Science, RDA, Iksan 570-080; National Academy of Agricultural Science, RDA, Suwon 441-407; Chonbuk National University, Jeonju 516-756 2

3

We developed duplex primer kit (Xoo-DP kit) for rice bacterial blight caused by Xanthomonas oryzae pv. Oryzae (Xoo) for rapid and accurate detection. The Xoo-DP kit was combination of Xoo-phage and Xoo-tnp primer. Xoo-DP kit were designed to specifically amplify a 402bp of int gene product phage-related integrase and 492bp of tnpA gene product transposase from X. oryzae pv. oryzae full genome sequence. Xoo-DP kit was used to carry out PCR analysis with genomic DNA from 25 different bacterial strains including phylogenetically related bacteria with X. oryzae pv. oryza, and other bacterial strains of different genus and species. The PCR assay using this XooDP kit was able to detect X. oryza pv. oryza with DNA concentration as low as 200fg and 3.7×10 cfu/ml of bacterial cell. The Xoo was detected from squashed rice leaf samples until extraction chlorophyll in 500ml of distilled water. 3

E-14

Downy Mildew on Perilla Leaf Caused by Pseudoperonospora cubensis in Korea. Chang Jeon Kim, Hye Yeon Mun, Je Yong Jung, Jin Pyo Hong and Hyang Burm Lee. Division of Applied Bioscience & Biotechnology, College of Agriculture & Life Sciences, Chonnam National University, Gwangju 500-757, Korea

Abstract of Presentations In the course of investigation of plant diseases on some crops, downy mildew was observed on perilla (Perilla frutescens L.) leaf in green houses located in Gwangyang, Korea. Further surveys revealed the disease to be widespread on this downy mildew species in other areas including Jeonbuk Province in Korea. On the basis of morphological characteristics and 18S and 28S rDNA sequences, the causal fungus was identified as Pseudoperonospora cubensis (Berk. et Curtis) Rostovtsev. To our knowledge, this is the first report of downy mildew on perilla caused by P. cubensis in Korea. The detailed morphological characteristics of the fungus will be presented in this poster.

E-15

Multiplex PCR method for the rapid simultaneous detection and diagnosis of the quarantine phytoplasmas. Young-Hwan Kim , 1

Chang-Ki Baek , Kyu-Ok Yim , Shigetou Namba and Hee-Young Jung . Division of Applied Biology and Chemistry, Kyungpook 1

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National University, Daegu, 702-701, Korea; International Quarantine Cooperation Division, National Plant Quarantine Service, Anyang 430-822, Korea; Department of Agricultural and Environmental Biology, The University of Tokyo, Tokyo 113-8657, Japan

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The homepage was also constructed to support for anyone to ask diagnosis pest on the cyber space. Main crops diagnosed in the web page were vegetable crops such as red pepper, cucumber, tomato etc. The total number of diagnostics was 60 from April to August including disease 20, insect 12, and physiological disorder 26. This system can be also a useful tool for collaboration and communication between extension agents and specialists to share information on plant diseases and pests.

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Survey of Phytophthora root rot of kiwifruit and screening of effective fungicides. Gyoung Hee Kim, Jin Hwi Moon, Kyoung

Youn Jo, Jae Sung Jung , Jae-Seoun Hur and Young Jin Koh. 1

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Department of Plant Medicine, Department of Biology and Department of Environmental Education, Sunchon National University, Suncheon 540-742, Korea 1

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The plant pathogenic bacteria not available in culture, phytoplasmas cause diseases of over one thousand plant species. Plant quarantine prevents the introduction and spread of phytoplasmas into the country, as well as certifying plants for export out of the country by PCR method. In this study, a PCR method for the rapid simultaneous detection and diagnosis of the quarantine phytoplasmas was constructed. On the basis of 16S rRNA gene phylogenetic analysis of all reported phytoplasmas, the results indicate that over 80 phytoplasma species exist in the world. However, only 13 species among them designated as the quarantine phytoplasmas in Korea. We have developed a PCR method for the rapid simultaneous detection and diagnosis of 10 species in the quarantine phytoplasma species. By multiple sequence alignments of several genes (16S rRNA, ribosomal protein, putative preprotein translocase, immunodominant protein, elongation factor TU and groEL), individual species-specific PCR primers designed from these unique sequences under consideration of some conditions. On the PCR performance, the species-specific PCR was able to identify phytoplasma species as appropriate length PCR products about 10 quarantine phytoplasma species. This PCR method will be used to rapidly and economically diagnose 10 phytoplasma species under same condition and only one reaction.

E-16

Construction of real time diagnosis network system in Gyeonggi province. Jin Young Kim, Soon Sung Hong, Jin Gu Lee, Hyun Ju

Lee, Young Hee Han and Sung Kee Kim. Gyeonggi-do Agricultural Research and Extension Services, Hwaseong 445-784, Korea

Outbreak of plant diseases and pests is now rapidly increasing because of global worming. So, the rapid monitoring and diagnostics of these became more important to reduce the spread of diseases and insects. In order to consult and diagnose of diseases and pests, real time network system was constructed including microscope, computer, server and web camera based on internet web page. This system enables client and county consultant to exchange the digital images and pictures and discuss the occurrence of disease and pest.

Phytophthora root rot has been severely occurred in major kiwifruit

cultivation areas in recent years. The overall incidence of the diseased trees infected by Phytophthora root rot was approximately 24% at the heavily damaged orchards in the major kiwifruit cultivation areas. The disease was relatively severe on the kiwifruit trees grown in paddy soils or poorly drained lowlands. Not only kiwifruit cv. ‘Hayward (Actinidia deliciosa)’ but also kiwifruit cvs. ‘Hort16A (A. chinensis)’ and ‘Hongyang (A. chinensis)’ were highly susceptible to the disease. Although the infected vines showed wilting and defoliation even on summer and severely infected trees resulted in death at late infection stage, no effective fungicide has been registered against Phytophthora root rot of kiwifruit in Korea until now. Twenty commercial fungicides were screened for their antagonistic efficacies based on the inhibition activities against mycelial growth of Phytophthora drechseri on 10% V8 juice agar. Mancozeb·metalaxyl WP showed the strongest inhibition activities, followed by copper oxychloride·metalaxyl WP, metalaxyl-M WP, dimethomorph· pyraclostrobin WG, dimethomorph WP, fluopicolide·propamocarb hydrochloride SC, chlorothalonil·oxadixyl WP. The antagonistic efficacies of mancozeb·metalaxyl WP, dimethomorph·pyraclostrobin WG and dimethomorph WP lasted during 12 incubation days on 10% V8 juice agar, but those of the other ones drastically decreased in proportion to incubation time. Field evaluations of the selected fungicides against Phytophthora root rot are still undergoing on kiwifruit orchards.

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Investigations of diseases and insect pests and characteristics of pear fruits wrapped with small bags. Young Jin Koh , Hong Chul 1

Kim , Kyoung Youn Jo , Gyoung Hee Kim , Jang-Hoon Song and Young-Sik Cho . Department of Plant Medicine, Sunchon National 1

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University, Suncheon 540-742, Korea; Pear Experiment Station, National Institute of Horticultural & Herbal Science, R.D.A, Naju 520-821, Korea 2

In order to confirm the possibility of cultivation of oriental pears without bags, the pear fruits wrapped with small bags on 25 May, 2009 were compared with those wrapped with common paper bags. Pesticides including systemic fungicides and insecticides for pear psylla (Psylla pyricola) and comstock mealybug (Pseudococcus comstocki) were sprayed 7 times at 10- to 15-day-intervals and Confuser-

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R○ was treated 2 times on early April and early August in order to control oriental fruit moth (Grapholita molesta) on the orchards. Occurrences of scab (Venturia nashicola) and cedar apple rust (Gymnosporangium asiaticum) of ‘Kosui’ and ‘Wonwhang’ were not affected by the kinds of bags. Incidences of skin sooty dapple (Cladosporium spp.) and sooty mold (Capnophaeum fuliginodes) were 19.2~24.6% on the fruits wrapped with common paper bags, while those were 29.3~53.3% on the fruits wrapped with small bags. Incidences of bird-injuries were 4.5~8.3% on the fruits wrapped with small bags, which were relatively high compared with those wrapped with common paper bags. Incidences of red fruits were 17.2% on ‘Wonwhang’ of which size is bigger than ‘Kosui’. The soluble sugar contents of fruits of ‘Kosui’ and ‘Wonwhang’ wrapped with small bags were higher than those wrapped with common paper bags but the acidity of fruits of ‘Kosui’ and ‘Wonwhang’ wrapped with small bags was lower than those wrapped with common paper bags. This reveals that the taste of fruits of ‘Kosui’ and ‘Wonwhang’ wrapped with small bags is better than that wrapped with common paper bags. L-value of ‘Kosui’ wrapped with small bags was 48.37 but that wrapped with common paper bags was 54.38, while a-value of ‘Kosui’ wrapped with small bags was 0.81, which was relatively high compared with that wrapped with common paper bags. r

F-3

Chemical control of scab of Japanese apricot and screening of resistant cultivars. Jong Sup Shin , Won-Ki Lee, Han Ju Park, Hyun

Jin-Kyoung Kim*, Chang-Ki Shim , Sang-Won Park , Byung-Jun Park, Won-Il Kim, Hyeong-Jin Jee , Oh-Kyung Kwon and Geon-Jae Im. Hazardous Chemistry Division, Organic Agriculture Division, 3

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National Academy of Agricultural Science, RDA; Research Policy Bereau., RDA; Agrobiodiversity Center, Rural Development of Administration, Swon 441-853, Korea 2

3

This study was conducted to develop an organic control of powdery mildews of cucumber by using mayonnaise in green house. The treatment of 0.1~2% mayonnaise resulted in 8.3%~99.2% control efficiencies against powdery mildew of cucumber. 0.5% mayonnaise treatment resulted control values over 97% in disease. It did not adversely affect the photosynthesis of foliages. Although one application of mayonnaise to the foliage was not practically effective enough, two or three application of mayonnaise to the foliage at the 0.5% concentration resulted in excellent control against powdery mildew. This treatment could provide protection for 10~14 days after application. Among the type of mayonnaise, general mayonnaise revealed 97.5% control value, but mayonnaise of low oil content revealed 39.3%~97.5% control values on powdery mildews at the 0.5% concentration. Oil content in mayonnaise played an essential material to control powdery mildew. Results indicated that mayonnaise could be used as organic control of powdery mildews of cucumber. This control might be environment-friendly as well as costeffective.

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Su Park , Kyoung Youn Jo , Gyoung Hee Kim , Myoung Taek Lim , Gil Ho Shin and Young Jin Koh . Suncheon-Si Agricultural Technol1

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ogy and Extension Center, Suncheon 540-804, Korea; Department of Plant Medicine, Sunchon National University, Suncheon 540-742, Korea; Jeonnam Agricultural Research and Extension Service, Naju 520-715, Korea 2

2

Occurrence of scab disease on Japanese apricot (Prunus mume) was surveyed at 45 orchards randomly selected among the major cultivation areas of Jeonnam and Gyeongnam in Korea during the 2009 growing season. The overall disease incidence of scab averaged 25.2%, but the incidences of scab ranged from 0% to 100% varied with the management conditions of the orchards surveyed. Since outbreak of scab disease at harvest stage resulted in heavy economic losses due to poor quality of the diseased fruits of Japanese apricot, it is important to prevent the epidemics before harvest. Control of scab has mainly depended on 1 to 3 times spray of fungicides at farmers' orchards. Incidences of scab were less than 22% at the sprayed orchards, while those of scab ranged from 56% to 100% were observed at the unsprayed ones. Trifloxystobin WG, dithianon WP, fenbuconazole WP and azoxystrobin+ difenoconazole SC were selected as effective fungicides against scab of Japanese apricot. Trifloxystobin WG sprayed two times at 10-day intervals from mid April exhibited more than 96% of control efficacy. Twenty-two cultivars of Japanese Apricot were screened for their resistance against scab by artificial inoculation of spore suspensions of Cladosporium carpophilum on the detached fruits in the laboratory. Japanese Apricot cv. ‘Okyoung’ was turned out to be resistant to scab.

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Control of Powdery Mildews of Cucumber by Using Mayonnaise.

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Environmental-friendly disease management of powdery mildew and leaf mold of tomato. Eun-Jung Choi , Kyung-Ryul Ryu , Seung1

2

Don Lee , Jong-Ho Park , Sung-Jun Hong , Hyeong-Jin Jee and Yong-Ki Kim . Organic Farming Technology Division, National 3

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Institute of Agricultural Science and Technology(NIAST), Rural Development Administration(RDA), Suwon 441-707, Korea; Research Planning Division, R search Policy Bureau, RDA, Suwon 441-707, Korea; Research Coordination Division, R search Policy Bureau, RDA, Suwon 441-707, Korea 2

3

Powdery mildew and leaf mold were major diseases in organic cultured tomatoes. NaHCO , Ca(NO ) and KH PO were selected as organic materials for controlling tomato powdery mildew. Control effect of the selected organic materials was increased when they were treated with oil-egg yolk mixtures (OEYO). Four organic materials used commercially including copper hydroxide and sulfur, showed high control effect more than 90% in green house. Also two organic matters, copper hydroxide and sulfur, showed high control effect in the farmer's field condition. When tomatoes were cultivated in plastic house installed with circulation fan, incidence of powdery mildew and leaf mold was reduced by 56% and 60%, respectively. Also leaf mold and powdery mildew were occurred less on ‘Super-top’ than ‘Totaerang’. 3

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Field study for anthracnose validation of an integrated pest management system for pepper at Bongwha and Taean in 2009. Sung-Taek Kim, Jeong-Wook Shin and Sung-Chul Yun. Depart-

ment of Biomedical Sciences, Sun Moon University, Asan 330-744, Korea

Abstract of Presentations Field study for validation of an integrated pest management (IPM) for pepper Phytophthora blight, tobacco budworm, and anthracnose was conducted 8 commercial fields at Bongwha, Kyungbuk and Taean, Chungnam in 2009. Five fields at Bongwha and three fields at Taean were cultivated by the owner and each field had three treatments: 1) No pesticide spray, 2) Model-advised spray, and 3) Conventional spray by farmer. The all eight fields were within 3 Km from the fieldweather stations in each location. Cultivar Burumi and Sangkang were cultured at Bongwha, and cvs. Umchungnan and Muhanjilju were cultured at Taean. The model-advised sprays were five times at the two locations, whereas the eight famers had sprayed 8-15 times at their conventional plots. At the end of the season at Bongwha, the fresh weight of red pepper yield, i.e. kg/40 plants, at no spray control were 21.4 kg of Burumi and 19.6 kg of Sangkang, whereas those at the model spray were 28.6 kg and 27.1 kg and those at the conventional spray were 29.9 kg and 31.7 kg, respectively. On the other hands at Taean, the red-pepper yield were 4.6 kg of Umchungnan and 9.6 kg of Muhanjilju, whereas those at the model spray were 8.8 kg and 15.2 kg and those at the conventional spray were 16.7 kg and 17.7 kg, respectively. Anthracnose model seemed to be acceptable if the anthracnose was not significantly severe at field, i.e. cvs. Brumi and Sangkang at Bongwha and cv. Muhanjilju at Taean, because it lost 4%, 15% and 14% of red pepper yield compared to the conventional spray, respectively. However, the forecaster did not work well for cv. Umchungnan at Taean where anthracnose was severe at no spray control as 80%, because it lost 47% of red pepper yield compared to the conventional spray.

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Influence of Late Blight Caused by Phytophthora infestans on Yield of the Potato. Jeom-Soon Kim , Young-Gyu Lee , Kyeong1

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Hun Park and Soon-Pil Kang . Highland Agriculture Research Cen2

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ter, National Institute of Crop Science, Rural Development Administration, Pyeongchang 232-955, Korea; National Academy of Agricultural Science, Rural Development Administration, Suwon 441-407, Korea 2

Late blight of potato, caused by Phytophthora infestans, has the potential to be a very destructive disease and to cause the tremendous yield loss of potato in Korea, especially in highland area. This study was conducted to find out the influence of late blight on potato yield with three potato cultivars. The initial disease severity ranged from 0 to 19.6% and from 0 to 5% in susceptible cultivars ‘Superior’ and ‘Atlantic’, respectively and from 0 to 5% in a slightly resistant cultivar ‘Haryeong’. The initial disease severity showed positive correlations with the area under the disease progress curves (AUDPC) and negative correlations with total and marketable yield in three cultivars. And correlations between disease severities and the total yields were the most apparent in those at 78 days after planting in three cultivars (Superior: y = -18.43x + 4143.8, R = 0.98, Atlantic: y = -10.31x + 3375.3, R = 0.97, Haryeong: y = -24.67x + 4467.3, R = 0.99). 2

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Suppression Effect of Postharvest Decay of Pear by Sterilization of Storage Box Using Volatile Natural Compound and Easy Materials. Ju-Hee Kim , Seong-Soo Cheong , In-Yong Choi , Dong1

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Chil Choi , Mi-Sun Baek and Wang Hyu Lee . Jeollabuk-do Agri1

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cultural Research and Extension Services, Iksan 570-704, Korea; Faculty Biological Resources Science, Chonbuk National University, Jeonju 561-756, Korea 2

In our previous studies, we have obtained antifungal inhibition activity of acetic acid and allyisothiocyanate for pathogen isolated from postharvest pear decay. This study was carried out to find control method of postharvest pear decay for applied to the field. The materials that is used at reduced to disease incidence were acetic acid and allyisothiocyanate having antifungal activity, sodium hyphochlorite, and chitosan etc. Allyl isothiocyanate fumigation inhibited up to 70.8, 72.5, 79.3% disease incidence of pear decay during storage by Penicillium spp., Colletotrichum sp. and Botrytis cinerea with concentration of 1.2 ml per m for 24, 48, 72hr, respectively. Acetic acid fumigation inhibited up to 74.3, 66.8, and 75.8% disease incidence of pear decay during storage by Penicillium spp., Colletotrichum sp. and Botrytis cinerea with concentration of 200µl of 4% per box for 24, 48, and 72hr, respectively. Sodium hyphochlorite wiping after washed and non-washed the box inhibited up to 20% and 48.5% disease incidence of pear decay during storage by pathogens with concentration of 200ppm per box, respectively. Chitosan wiping after washed and non-washed the box inhibited up to 34 and 48.5% disease incidence of pear decay during storage by pathogens with concentration of 200fold, respectively. The pears treated with materials treatment for storage box after washing were found to be more effective in reducing disease severity than non-washing box. These results indicated that acetic acid, allyisothiocyanate, and sodium hypochlorite should have value to use as a suppression agent for controlling postharvest pear disease. 3

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Studies on the efficacy of soil amendments with neem cakes and with bio-control agent on the incidence of Macrophomina stem and root rot of sesame. T.S. Rajpurohit and S. Dubal. Agricultural Research Station, Rajasthan Agricultural University Mandor, Jodhpur- 342 304 (Raj.) India

Stem and root rot of sesame caused by Macrophomina phaseolina infects high percentage of plants and consequently leads to great yield losses in rainfed crop especially in Rajasthan. The continuous use of chemicals has deleterious effect on the beneficial microorganism in soil, in addition to the residual problem and development of resistance by the pathogen. An experiment was conducted in randomized block design with eight treatments and four replications with plot size of 4×2.4m. on sesame during kharif 2006 - 2007 at Agricultural Research Station, Mandor - Jodhpur (Rajasthan) - India to find out the efficacy of soil amendments with neem cakes and with bio-control agent on the incidence of Macrophomina stem and root rot. The cakes were incorporated in soil and mixed thoroughly before sowing. Bio agent Trichoderma viride was added in FYM 15 days prior to its application and kept in shed and the incidence of Macrophomina stem and root rot was recorded before harvesting. Minimum incidence of Macrophomina stem & root rot (3.32%) and highest seed yield (924 Kg/ha) was recorded in soil application of Neem cake (250 kg/ha) + seed treatment with Trichoderma viride (0.4%) + soil application of T. viride @ 2.5 kg/ha, This treatment gave 82.27% disease control and 43.92 % yield increase with B : C ratio of 2.88, this was followed seed treatment with T. viride (0.4%) + soil application of T. viride @

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2.5 kg/ha (PDI 6.08%, seed yield 816 kg/ha). Highest disease (13.03%) was recorded in control. Two years results revealed that soil application of Neem cake( 250 kg/ha) + seed treatment with Trichoderma viride (0.4%) + soil application of T. viride @ 2.5 kg/ha reduced Macrophomina stem & root rot from 18.17 to 3.32% and increased seed yield from 642 kg/ha to 924 kg/ha with B : C ratio of 2.88.

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Okabe1 and Koya Sugawara. National Institute of Livestock and

Plant growth promoting microbial consortia mediated biological control of sunflower necrosis virus disease under natural field conditions. K. Srinivasan* and N. Mathivanan. Biocontrol and

plots covered with lentil tare were lower than in plots mulched with black vinyl. Actinomycetes, bacteria, and fungal densities were significantly greater in rhizosphere soil in plots covered with lentil tare compared with soil in black vinyl mulching.

Resistance of Italian ryegrass to blast disease and fitness of the pathogen on the host. Takao Tsukiboshi, Takako Kiyoshi, Ikuko Grassland Science, Nasushiobara 329-2793, Japan

Sunflower necrosis virus disease (SNVD) is severely affecting sunflower cultivation in India with disease incidence up to 90%. Biological control is the best option for managing SNVD as no other effective methods are available. In this study, two formulations (powder & liquid) of Bacillus licheniformis MML2501 (Bl), Streptomyces fradiae MML1042 (Sf), Pseudomonas aeruginosa MML2212 (Pa) and Bacillus sp. MML2551 (Bsp) were developed individually and evaluated as three (Bl + Pa + Bsp) and four cultures (Bl + Sf + Pa + Bsp) consortia against natural SNVD in the farmers field. These consortial formulations remarkably enhanced the plant growth and reduced SNVD there by increasing the seed yield. SNVD reduction was estimated up to 51.4% and 40.9%, due to four cultures consortium of liquid and powder, respectively formulations compared to control. Further, it significantly increased the yield parameters than the other treatments. Liquid formulation of four cultures consortium registered an additional seed yield of 936 kg/ha with an additional income of Rs. 12168/ha whereas, talc formulation registered an additional seed yield 840 kg/ha with an additional income of Rs. 10920/ha compared to control. The results of field experiments clearly demonstrated that the both the four cultures consortium formulations consisting of Bl + Sf + Pa + Bsp can be used for the effective control of SNVD.

Blast caused by Magnaporthe oryzae is one of the most important diseases of Italian ryegrass, a grass species widely used as forage and turf in temperate and subarctic region, including Japan. Its damage is especially severe on susceptible cultivars under warm and wet weather. Since most of the cultivars are susceptible to the disease, breeding of resistant cultivars are of the interest in the countries cultivating the grass. To observe the cytological reaction of resistant breeding lines to the disease and estimate the fitness of the pathogen, inoculation tests have done under different environmental conditions. Selected resistant lines of Italian ryegrass, ‘Yamaiku 180’ and ‘Yamaiku 181’, showed strong resistance to the disease producing no visible lesions after inoculation. Papilla-like structures were produced in the plant cells just beneath the appressoria of the pathogen and likely to be preventing its invasion. However, no hypersensitive cell death was observed. A resistant cultivar, ‘Sachiaoba’, the only resistant cultivar currently available in Japan, showed resistance to produce few and small lesions on the infected plants in fields. The pathogen showed lower fitness on the cultivar producing smaller number of conidia per lesion due to suppressed enlargement of the lesions compared to susceptible cultivars. Number of conidia per lesion was also affected by temperatures and was highest at 28-31°C compared to lower temperatures of 19-22°C. Higher fitness of the pathogen seen under higher temperature might have some affect on the epidemic and expanding distribution of the disease in Japan under global warming situation.

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Microbial Metabolites Lab, Centre for Advanced Studies in Botany, University of Madras, Guindy Campus, Chennai - 600 025, India

Effect of living mulching by lentil tare on pepper field. Young Kee

Lee , Jae Hyeon Park , Jung Lai Cho , Inn Shik Myung , Jae Kook Lee and Sang Yeob Lee . Agricultural Microbiology Division, 1

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National Academy of Agricultural Science (NAAS), Rural Development Administration (RDA), Suwon 441-707, Korea; Organic Agriculture Division, NAAS, RDA, Suwon 441-707, Korea 2

This study was conducted to examine the influence of pepper (Capsicum annuum L.) with two mulches on occurrence of anthracnose and bacterial spot, growth of the plant, and microorganism densities in rhizosphere soil. Pepper was grown in bare ground or intercropped with lentil tare (Vicia tetrasperma (L.) Schreb.) or black vinyl. Occurrence of anthracnose on fruits of the pepper was less in plots covered with living mulch than in bare ground and plots mulched with black vinyl. Occurrence of bacterial spot on leaves of the pepper was higher in plots covered with lentil tare than bare ground, but no difference with plots mulched with black vinyl. Height of pepper in living mulching plots was similar in black vinyl mulching, but diameter of lower stem, number of fruit per plant, and length of fruit on pepper in

Anti-microbal activity of avenacin saponin to fungal pathogens for cereals. Yoshishige Inagaki , Atsuko Imaoka , Keiko Fujita , 1

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Sakae Arase , Anne Osbourn , Kazuhiro Toyoda , Tomonori Shiraishi and Yuki Ichinose . Graduate School of Natural Science 2

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and Technology, Okayama University, Tsushima-naka 1-1-1, Okayama 700-8530, Japan; Laboratory of Plant Pathology, Faculty of Life and Environmental Science, Shimane University, Matsue 6908504, Japan; John Innes Centre, Colney Lane, Norwich NR4 7UH, UK 2

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Triterpenes are natural products that, like sterols, are synthesized from 2,3-oxidosqualene. Simple triterpenes have been identified in the leaf waxes of various plant species. Some plants also produce elaborate triterpene glycosides (known as saponins). Triterpene glycosides are commonly anti-fungal (unlike simple triterpenes) and have been implicated in plant defense. Oat (Avena) is unusual amongst the cereals in that it produces anti-microbial triterpene glycosides. These rootderived compounds (avenacins) confer broad spectrum resistance to soil-borne diseases; Gaeunmannomyces graminis var. avenae, Fsar-

Abstract of Presentations ium anenaceum and so on (Mery et al, Phytochem. 25(9) 2069-2073,

1986). On the other hand, the importance of saponin detoxification in fungal pathogenesis has recently been demonstrated for some isolates of the fungus G. graminis var. avenae, which produces the enzyme avenacinase. Avenacinase detoxifies the triterpenoid oat root saponin avenacins, and is essential for pathogenicity of G. graminis var. avenae to oats. Here, to investigate the potentiality of avenacin for plant defense in other valuable cereals, we report fungicidal activity of avenacin to fungal pathogens; Magnaporthe grisea, Bipolaris oryzae, and Blumeria graminis f.sp. hordei, for other cereals (rice and barley).

G-3

Innate surface structures of pine needles as possible host defense mechanisms against fungal infection. Ki Woo Kim , In Jung Lee , 1

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Chang Soo Kim , Don Koo Lee and Eun Woo Park . National Instrumentation Center for Environmental Management, Seoul National University, Seoul 151-921, Korea; Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul 151921, Korea; Department of Forest Genetic Resources, Korea Forest Research Institute, Suwon 441-350, Korea; Department of Forest Sciences, Seoul National University, Seoul 151-921, Korea; Department of Agricultural Biotechnology, Seoul National University, Seoul 151921, Korea 3

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then oxidized by monoamine oxidase (MAO). As a consequence of this oxidation, H O is generated. In M. oryzae-infected leaves, TDC and MAO activities and the H O accumulation increased under light. DNA fragmentation and SL formation were induced by Try treatment under light, but not in the presence of the MAO inhibitor and H O scavenger. When M. grisea-infected leaves were kept under different visible light conditions (400~700nm), Try accumulation, TDC and MAO activities, and the H O generation were enhanced by increasing the wavelength of visible light. The effective wavelength for Try pathway activation agrees with that for induction of DNA fragmentation in leaves infected with M. oryzae. SL formation, Try accumulation and DNA fragmentation in leaves infected with M. oryzae were significantly inhibited even under light by pretreatment with cycloheximide and heat. Furthermore, glyphosate [N-(phosphonomethyl) glycine] suppressed SL formation and Try accumulation even under light. Such glyphosate inhibitions were blocked by the supply of exogenous tryptophan. Thus, we conclude that Try pathway plays an important role in light-dependent resistance of rice to M. oryzae infection. 2

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G-5

Role of a volatile in white-backed planthopper- induced resistance to bacterial blight in rice. Yumi Shinonaga , Masaru Satoh , 1

Epicuticular waxes and stomatal chambers were investigated in needles of Pinus rigida and P. densiflora. The two-year-old seedlings of the pine species were grown in a nursery and transferred to a growth chamber maintained at 25 C. Both immature and mature needles were collected and subjected to surface observations. Field emission scanning electron microscopy revealed that epicuticular wax structures developed on the cuticle layer and appeared to occlude stomata in the two pine species. The most prominent epicuticular wax structures of the two pine species included coiled rodlets and tubules. Either rodlets or tubules appeared to be longer and form less tight clogging in P. rigida than those in P. densiflora. The stomata of P. densiflora were characterized by more distinctly raised Florin rings than P. rigida. In vertical profiles by white light scanning interferometry, the stomatal chambers of P. densiflora were ca. 20 µm deep, whereas those of P. rigida were ca. 10 µm deep. These results suggest that such innate surface structures of the pine species could contribute to different levels of hyphal growth to natural openings on the needle surface.

G-4

Role of trypatmine pathway in light-dependent resistance of rice to Magnaporthe oryzae infection. Makoto Ueno, Yumiko Kumura, Kana

Ueda, Atsuko Imaoka, Junichi Kihara, Sakae Arase, Akinobu Oshima and Keisuke Yoshikiyo. Faculty of Life and Environmental Science, Shimane University, 1060 Nishikawatsu, Mastue 690-8504, Japan

Lesion mimic mutant of rice (cv. Sekiguchi-asahi) showed lightdependent resistance to Magnaporthe oryzae infection inducing the Sekiguchi lesion (SL) formation. The hyphal growth and sporulation are not observed in the SLs. In leaves with SL, tryptamine (Try) accumulation and DNA fragmentation are induced. SL formation, Try accumulation and DNA fragmentation were also induced by treatment with the spore germination fluids from M. oryzae. Try is biosynthesized from tryptophan by tryptophan decarboxylase (TDC), and

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Rika Ozawa , Hiroo Kanno , Jyunji Takabayashi , Kazuya Akimitsu and Kenji Gomi . Department of Life Sciences, Faculty of Agricul3

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ture, Kagawa University, 761-0795, Miki, Kagawa, Japan; National Agricultural Research Center for Kyushu Okinawa Region, NARO, Koshi, Kumamoto, 861-1192, Japan; Center for Ecological Research, Kyoto University, Otsu, Shiga, 520-2113, Japan 2

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Interspecific interactions between organisms utilizing the same plant have been one of the important subjects in community ecology. Recently, we found that resistance to rice blast caused by Magnaporthe grisea in rice is induced by white-backed planthopper (WBPH), Sogatella furcifera Horváth, one of the phloem-sucking insects to rice plants. WBPH infestation also conferred resistance to Xanthomonas oryzae pv. oryzae (Xoo), which causes a vascular wilt disease, known as ‘bacterial blight’, one of the most serious rice-plant diseases in rice growing countries. The infestation of another planthopper species, the brown planthopper (BPH) Nilaparvata lugens Stål, did not induce the significant resistance to Xoo in rice. By GC/MS analysis, it was revealed that one monoterpene, linalool, accumulated in rice after WBPH infestation but not after BPH infestation. WBPH infestation caused the upregulation of linalool synthase gene than did BPH infestation. A vapor treatment of linalool to rice induced resistance to Xoo. A direct treatment of linalool to a liquid culture of Xoo had no effect on the growth of Xoo, indicating that linalool has no antibacterial activity to Xoo. These results strongly indicate a possibility that linalool plays an important role in WBPH-induced resistance as a signal compound in rice.

G-6

The selection of desirable cultivar for organic cultivation of carrot. Sang Jun Lee, Jae Eun Lee, Eun Kyung Jung, Zhang Yan, Cheol Soo Yoon, Young Rok Yong and Byung Sup Kim. Department of Plant Science, Gangneung-Wonju National University, Gangneung 210-702, Korea

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This research was carried out to selection of desirable cultivar for organic cultivation of carrot in Korea. The assay of disease resistance, insect tolerance and rate of bolting were investigated on 33 cultivars of genetic resources of carrot. This experiment was carried out at experimental fields of Gangneung-Wonju National University. “PI 223360” was resistance against powdery mildew. Ten cultivars including “PI451752” were moderate resistance, 22cultivars including “Long Impeator #58” were susceptible to powdery mildew. Although there were no cultivars against black rot and leaf blight, our results were confirmed that several cultivars have moderate resistance. Thirteen cultivars including “PI451752” were having insect tolerance. According to the investigation of bolting, 6 cultivars including ‘Flavis’ were early bolting and 4 cultivars including “NutriRed” were moderate bolting. Other cultivars were identified as late bolting. From above results, we confirmed that “Oxheart carrot heirloom” was suitable cultivar for organic cultivation among 33 genetic resources of carrot.

identified tobamovirus resistance gene that was mapped to the L locus, confers resistance against the tobamovirus P0 pathotype at 30°C. We cloned the L gene from Capsicum annuum KC780 using PCR with a primer set designed based on the sequence of L gene, which was cloned by a map-based method. By using the Agrobacterium-mediated transient expression assay in Nicotiana benthamiana leaves, we analyzed the functional domain in L and tobamovirus effecter coat protein (CP) for induction of hypersensitive response (HR). Co-expression of L and CP of Tobacco mild green mosaic virus (TMGMV, P0 pathotype) resulted in HR at 30°C. By contrast, HR was not obviously observed when L and TMGMV-CP were coexpressed at 30°C. Six amino acid changes were observed between L and L . Chimeric L , in which two amino acid residues in putative LRR domain were replaced with the equivalents of L , induced weaker HR than original L at 30°C. To analyze the functional domain of TMGMV-CP for elicitation of HR at high temperature, chimeric CP between TMGMV-CP and Paprika mild mottle virus (PaMMV, P1 pathotype) CP were constructed. When co-expressed with L , TMGMV-CP obviously elicited HR both at 24°C and 30°C. On the other hand, PaMMV-CP obviously elicited HR at 24°C, but not at 30°C. Chimeric TMGMV-CP, in which 22 amino acids (residues 50 to 71) region was replaced with the equivalent of PaMMVCP, obviously elicited HR at 24°C, but not at 30°C (identical to that of PaMMV-CP). These observations indicate that two amino acid residues in putative LRR domain in L and 22 amino acid residues in TMGMV-CP are involved in the ability of HR induction by L at high temperature. 1a

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G-7

Resistant Cultivar screening to clubroot (Plasmaodiophora brassicae) for cultivation of Chinese cabbage in Gangwon alpine area. Cheol Soo Yoon, Eun Kyung Jung, Lee Sang Jun, Zhang Yan, Jae Eun Lee and Byung Sup Kim. Department of Plant Science, Gangneung-Wonju National University, Gangneung 210-702, Korea

1a

1a

This study was conducted to screen the cultivar which is suitable in cultivation of the Chinese cabbage having good horticultural characteristics and the resistance to clubroot (Plasmaodiophora brassicae) caused serious problems in many Brassicaceae growing areas. Fifty three cultivars of the Chinese cabbage were planted in the fields contaminated with clubroot pathogen (Race 8) and investigated the disease occurrence rate and severity of clubroot and the horticultural characteristics. In case of above-ground rate of clubroot, cv. Dae-Tong, cv. Bulamplus, cv. super CR Hwang-Mi 85, cv. super CR Hiroki, cv. SP-05, cv. SP-02, cv. Gaenari, cv. CR DB and cv. CR DB 29 were resistant to clubroot in 30 days after transplanting. In case of below-ground clubroot rate, cv. Dae-Tong, cv. super CR Hwang-Mi 85 and cv. super CR Hiroki were resistance to clubroot in 60 days after transplanting. In comparison of fresh weight of Chinese cabbage among cultivars, cv. super CR Hiroki as over 2,500g was the best than any other cultivars. From above the results, we recommend that super CR Hiroki will be more suitable in cultivation of Chinese cabbage in alpine area.

G-8

Analysis of the functional domains in Capsicum plant resistance protein L and Tobacco mild green mosaic virus effecter coat protein for HR induction at high temperature. Hiroyuki Mizumoto , 1a

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Ikumi Nakamura , Akinori Kiba , Reiko Tomita , Kappei Kobayashi and Yasufumi Hikichi . Laboratory of Plant Pathology and Biotech1

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nology, Kochi University, 200 Monobe, Nankoku, Kochi 783-8502, Japan; Iwate Biotechnology Research Center, Kitakami, Iwate 0240003, Japan

G-9

Evaluating Resistance Against Anthracnose in Grapevines by Treating Culture Filtrates from Pathogen, Elsinoe ampelina. Hae Keun Yun *, Myung Hwan Jang , Seung Hui Kim and Jeong Ho Noh . Department of Horticultural Science, Yeungnam University, 1

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Gyeongsan 712-749, Korea; Fruit Research Division, National Institute of Horticultural and Herbal Science, RDA, Suwon 440-706, Korea 2

As the control of anthracnose occurring severely in grapevines is very difficult, resistance of some cultivars to anthracnose is one of the big advantages in grapevine cultivation. It was investigated whether culture filtrates produced by Elsinoe ampelina could be used to determine varietal susceptibility in grapevines to anthracnose as a substitute for pathogen inoculation or field screening. Bioassay of grapevine leaves with culture filtrate and its ethyl acetate extracts showed that their phytotoxicities were active and host-selective. In the result of varietal resistance evaluation by treating culture filtrates among grapevine cultivars, some such as ‘Black Swan’, Rizamat’, ‘Rosario Bianco’, and ‘Kaiji’ were sensitive, and others such as ‘Campbell Early’, ‘Niagara’, ‘Sheridan’, and ‘Izumo Queen’ were found to be tolerant to anthracnose. Evaluation for resistance in the vineyard showed the consistent results with those obtained from bioassay with culture filtrates of the pathogen.

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Tobamovirus resistance in Capsicum plants, which is confered by L genes (L , L , L or L ), is known to be compromised under high temperature conditions, above 30°C. However, the L gene, a newly 1

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Profiling of ESTs Related with Defense Responses from Vitis coignetiae. Hae-Keun Yun , Youn-Jung Choi and Youn-Young Hur . 1

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Department of Horticultural Science, Yeungnam University, Gyeong-

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san 712-749, Korea; Grape Research Center, National Institute of Horticultural and Herbal Science, RDA, Suwon 440-706, Korea

G-12

Vitis coignetiae, a kind of Korean domestic wild grapes, have been

Akihito Kano1, Kouji Hosotani , Takeshi Fukumoto , Kouhei Ohtani , Ken Izumori , Shigeyuki Tajima , Toshiaki Ohara , Yoshio Shigematsu , Keiji Tanaka , Yutaka Ishida , Kenji Gomi , Yasuomi Tada and Kazuya Akimitsu . Faculty of Agriculture and Gene

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known to be resistant to diseases. For screening useful resources for disease resistant grape breeding, cDNA library was constructed from matured leaves and fruits of wild grapes. A total of 2,982 unigenes containing 676 contigs and 2,306 singletons were obtained by sequencing 5,760 expressed sequence tag (EST) clones derived from cDNA library of wild grapes. In gene ontology analysis, 2,241 molecular function, 1,442 biological process, and 836 cellular component genes were annotated. Defense-related genes such as putative prolinerich cell wall protein, thaumatin-like protein, class IV chitinase, and pathogenesis-related protein 10, and photosynthesis-related genes such as ribulose-bisphosphate carboxylase oxygenase small subunit, and water stress-related genes such as abscisic stress ripening protein and aquaporin were the most abundant in cDNAs from grapes and leaves of V. coignetia. These data could provide useful information in genetic analysis of V. coignetia and in the program for breeding disease resistant grape cultivars.

G-11

Airborne induction and priming of plant defenses against a bacterial pathogen. Hwe-Su Yi , Martin Heil , Rosa M. Adame1,2

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Álvarez , Daniel J. Ballhorn and Choong-Min Ryu . Laboratory of 3

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Microbial Genomics, Industrial Biotechnology and Bioenergy Research Center, KRIBB, Daejeon 305-806, Korea; School of life science, Kyungpook National University, Daegu 702-701, Korea; Departamento de Ingeniería Genética, CINVESTAV Irapuato, 36821 Irapuato, Guanajuato, Mexico; Dept. of General Botany-Plant Ecology, University of Duisburg-Essen, Essen, Germany; Field of Functional Genomics, School of Science, University of Science and Technology (UST), Daejeon 305-333, Korea 2

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Herbivore-induced plant volatiles affect the systemic response of plants to local damage and hence represent a kind of plant hormone. These signals are transported outside the plants and can, therewith, also lead to ‘plant-plant communication’, a defense induction in yet undamaged plants that grow close to damaged neighbors. We observed a similar phenomenon in the context of disease resistance. Lima bean (Phaseolus lunatus) plants became more resistant against a bacterial pathogen, Pseudomonas syringae pv. syringae, when they were located close to conspecific neighbors in which systemic acquired resistance (SAR) to pathogens had been chemically induced with benzothiadiazole (BTH) or biologically by infection with avirulent P. syringae. Challenge inoculation after exposure to induced and non-induced plants revealed that the air coming from induced plants mainly primed resistance, since expression of pathogenesis-related protein 2 (PR-2) was significantly stronger in exposed than in nonexposed individuals when the plants were subsequently challenged by P. syringae. Among others, the plant-derived volatile, nonanal, was present in the headspace of BTH-treated plants and significantly enhanced PR-2 expression in the exposed plants. Negative effects on growth of BTH-treated plants, which usually occur as a consequence of high costs of direct resistance induction, were not observed in VOC-exposed plants. Volatile mediated priming appears to be a highly attractive means for the tailoring of SAR against plant pathogens.

A rare sugar, D-allose, confers resistance to rice bacterial blight with accumulation of reactive oxygen species in Oryza sativa L. 1

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Research Center, Kagawa University, Miki, Kagawa 761-0795, Japan; Mitsui Chemicals Agro, Inc., Yasu, Shiga 520-2342 Japan; Sikoku Research Institute, Inc., Takamatsu, Kagawa 761-0192 Japan 2

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We investigated responses of rice plant to some of rare sugars. Rice growth was significantly inhibited by D-allose but not by the other rare sugars tested. A large-scale gene expression analysis using a rice microarray revealed that D-allose treatment causes a high upregulation of many defense-related PR-protein genes in rice. The PR-protein genes were not upregulated by other tested rare sugars. Furthermore, D-allose treatment of rice plants conferred resistance of rice against bacterial pathogen of Xanthomonas oryzae pv. oryzae (Xoo), but the other tested sugars did not. We also found that D-allose has no direct effect, such as an anti-bacterial activity to Xoo. These results suggested that D-allose confers resistant to bacterial blight by inducing the uncharacterized defense system(s) in rice. We also found that D-allose-treated rice exhibited a lesion mimics on the leaves. It has been reported that the lesion mimic following hypersensitive response was caused by production of reactive oxygen species (ROS) and this response associated with disease resistance to Xoo in rice. Thus, we performed detection of ROS in leaf blades. As the result, Dallose-treated rice leaf blades were accumulated with ROS higher than those of control and D-glucose-treated plants. In addition, Dallose-mediated induction of resistance to rice bacterial blight was inhibited by reducing agents such as ascorbic acid. These results indicate that the ROS accumulation by D-allose is involved in disease resistance to Xoo in rice.

G-13

Insight into non-host resistance against Ralstonia solanacearum on Nicotiana benthamiana. Hyo Bee Park and Choong-Min Ryu.

Laboratory of Microbial Genomics, Industrial Biotechnology and Bioenergy Research Center, KRIBB, Daejeon 306-805, Korea

Ralstonia solanacearum, a casual pathogen of bacterial wilt is eco-

nomically important pathogen in the field crops particularly solanaceae family. To date, no resistant plant variety has been reported against the pathogen. For establishing successful management, the nature of plant defense responses against R. solanacearum needs to be understood. Recent studies revealed that Nicotiana benthamiana showed clear resistance phenotype against certain strains of R. solanacearum indicating that N. benthamiana is a non-host plant species. To study mechanism(s) related to the non-host resistance against R. solanacearum, we employed Tobacco rattle virus (TRV) derived virus-induced gene silencing (VIGS) vector in Agrobacterium tumefaciens that have widely utilized on functional genomics of plant science. Two weeks after infiltration of TRV vectors, 106 colony forming units per ml bacterial suspension of R. solanacearum were drenched on the crown part of each silenced plants. Silencing of MATE, WIPK, WRKY, ZINC finger protein and reactive oxygen species related proteins in N. benthamiana displayed typical wilt

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symptoms on the leaves at two weeks after pathogen challenge. To examine the roles of these proteins in non-host resistance in N. benthamiana, we are now analyzing the corresponding gene silencing. Our results suggest that non-host resistance-related genes selected from our screening can be an important genetic resource to manage bacterial wilt disease in the agricultural crops.

G-14

The Kinds of Disease and Damage to Barley and Wheat in Korea.

Jong-Chul Park , Eun-Sook Lee , Chul-Soo Park , Chon-Sik Kang , Tae-Hwan Noh , Jung-Joon Lee , Jun-Hee Lee , Tae-Soo Kim , TaeIl, Park , Ki-Jong Kim and Jung-Gon Kim . Department of Rice and 1

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Winter Cereal Crop, NICS, Iksan 570-080, Korea; College of Agriculture & Life Science, Chonbuk National University, Jeonju 561756, Korea

gae pv. tomato DC3000 (Pst DC3000). Modulated cytokinin levels or signaling activity in CKX- or IPT-overexpressing plants or in ahk2 ahk3 lines correlated with resistance against Pst DC3000. Overexpression of cytokinin-activated transcription factor ARR2, but not ARR1, enhanced resistance to Pst DC3000. TGA3 specifically interacted with ARR2. Mutation of TGA-binding cis-elements in the PR1 promoter abolished cytokinin- and ARR2-dependent PR1 or PR2 activation. Moreover, applied cytokinins did not compromise the pathogen susceptibility of npr1-1 and NahG plants, as the cytokinindependent induction of PR1 was abolished. Taken together, cytokinins modulate the SA signaling cascade to augment resistance against Pst, and the interaction between TGA3 and ARR2 is important in this process.

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The symptom expressions such as yellowish and mosaic spots in overwintering barley had revealed by viruses. The major viruses were Barley yellow mosaic virus(BaYMV) and Barley mild mosaic virus(BaMMV) detected more than 70% and 15.7-37.4% in diseased barley leaves. Soil-borne wheat mosaic virus and Barley yellow dwarf virus also detected even it was a little ratio comparing to other viruses. BaYMV infection affected on cellular elongation or growth not cell division in examined three parts as stem, neck of panicle and node, related to dwarfness of infected barley. According to above cell damage resulted in shorten plant growth like as retarded culm length and the yield reduction over 70% in susceptible hulless barley cultivar. In 2009, we collected and investigation of other disease occurrence in barley and wheat fields. On kernel or head, discolored spot was major symptom and 3 kinds of pathogenic and other unidentified fungi was isolated such as Fusarium spp.(8%), Epicoccum spp.(60%), two kinds of Alternaria spp.(30-37%). On leaves and stem, blight, necrotic spot and stem dead were general symptoms. Sharp eyespot (Rhizoctonia cerealis) was identified and occurred in first in Korea and it caused yield reduction to 30% in wheat. Several pathogenic fungal and bacteria also isolated such as Pseudomonas syringae pv. syringae, Cladosporium spp. Alternaria spp. Nigrospora spp. Fusarium moniliforme. It need more accurate identification and continuous examination diseases occurrence and damage.

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G-16

Resistance Screening of Pepper (Capsicum annuum L.) Cultivars to Powdery Mildew (Leveillula taurica). Dae-Hyun Kim, Jung-Sup Lee, Jong-Han Park, Kyung-Sook Han, You-Kyoung Han, DooWook Kim and Jeong-Hwan Hwang. National Institute of Horticultural & Herbal Science, RDA. Suwon 441-440, Korea

The use of fungicides for the control of powdery mildew (Leveillula taurica) in pepper (Capsicum annuum L.) has been ineffective. Therefore genetic resistance is the best alternative. The purpose of this work was to evaluated for the resistance to powdery mildew in pepper cultivars. Fifty-one pepper cultivars including breeding lines in National Institute of Horticultural & Herbal Science, RDA were evaluated for the resistance to powdery mildew under field and green house, respectively. The tested pepper cultivars showed different degrees of resistance to powdery mildew. Disease severity ranged from 30 to 70%. Most cultivars (F ) were susceptibility to the powdery mildew, however, new breeding cultivars, AR 1, 2, 3 and 4 (C. baccatum×C. annuum) were moderately resistance. C. annuum materials known to be classified as moderately to highly susceptible to powdery mildew. Conversely, C. baccatum were most often resistant sources. Therefore, they will be recommended as resistant cultivars in greenhouses where powdery mildew are severe problem. 1

G-17

Wildfire (Pseudomonas syringae pv. tabaci) Resistance Screening of Soybean Cultivars in Field. Inn-Shik Myung , Jong Hyeong Lee , 1

Cytokinins promote plant immunity against a bacterial pathogen via ARR2-elicited TGA3 and NPR1-dependent salicylic acid signaling in Arabidopsis. Jaemyung Choi , Sung Un Huh , Mikiko 1

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Kojima , Hitoshi Sakakibara , Kyung-Hee Paek , Yongsoo Kim and Ildoo Hwang . Department of Life Sciences, Pohang University of 3

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Science and Technology, Pohang 790-784, Korea; School of Life Sciences and Biotechnology/Plant Signaling Network Research Center, Korea University, Seoul 136-701, Korea; Plant Science Center, RIKEN (The Institute of Physical and Chemical Research), 1-7-22, Suehiro, Tsurumi, Yokohama 230-0045, Japan

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Young-Kee Lee and Hong Sik Shim . Agricultural Microbiology 1

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Division, National Academy of Agricultural Science, Rural Development Administration, Suwon 441-707, Korea; Northern Agricultural Research Station, Gyeonggi Province Agricultural Research and Extension Services, Yeoncheon 486-833, Korea 2

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Many pathogenic bacteria and fungi produce cytokinins to enhance their pathogenicity; however, the role of plant-derived cytokinins in the pathogen response has been elusive. In this study, we found that cytokinins promote resistance of Arabidopsis to Pseudomonas syrin-

Eight varieties of soybean were screened for resistance to wildfire caused by Pseudomonas syringae pv. tabaci using spraying-inoculation technique in field. Diseased leaf area ranged from 0 to 50% over a severity scale of 0 (zero infection) to 5 (over 50% infection). Four cultivars of Shipaldal 2, Gumjeong 4, Daepung, and Chungja 3 were resistant. Three cultivars of Jangwon, Hwanggeum, and Daewon were susceptible. A cultivar of Nuetseoritae was moderate-resistant. The field screening technique was compared with a technique using sab-inoculation after cutting leaf with knife in greenhouse.


G-18

Differential interactions of Phytophthora capsici JHAI1-7 with hot pepper cultivars at various plant growth stages and inoculum concentrations. Su-jung Jo, Jin-Cheol Kim, Kyoung Soo Jang, Young Ho Choi and Gyung Ja Choi. Chemical Biotechnology

Research Center, Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea

Hot pepper cultivars were evaluated for resistance to Phytophthora blight caused by Phytophthora capsici JHAI1-7 under controlled environmental conditions. By using soil-drenching inoculation method, six cultivars of hot pepper (Bukang, PR-Sadaechunwang, PRDadda, Yeokganghongjanggun, PR-Power, and CM334) with different degrees of resistance to P. capsici JHAI1-7 were tested at various plant growth stages. As plants grew older, all cultivars became gradually resistant to Phytophthora blight. On the other hands, high resistant cultivars (PR-Power and CM334) showed highly resistant response to the pathogen at two-leaf stage of host seedlings as well as at more than six-leaf stage of hot pepper. At inoculum concentrations of 3×10 , 1×10 , and 3×10 sporangia/ml, resistance of seven cultivars (Bukang, PR-Sadaechunwang, PR-Dadda, Yeokganghongjanggun, Dokyacheongcheong, PR-Power, and Muhanjilju) to the pathogen was also assessed. The disease severity of all test cultivars was proportional to the inoculum concentrations. Bukang was a highly susceptible cultivar, PR-Sadechunhwang, PR-Dadda, Yeokganghongjanggun and Dokyacheongcheong were a moderate resistant, PR-Power, Dokyacheongcheong, Muhanjilju and CM334 were high resistant to the disease. So, our results suggest that screening pepper cultivars for resistance to P. capsici could be done effectively at 2-leaf stage of plants and by soil-drenching inoculation with zoospore suspension (3×10 or 1×10 sporangia/ml). 4

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G-19

Investigation of Phytophthora root rot resistance caused by Phytophthora capsici for evaluation of Tomato germplasm (Lycopersicon spp.). Jeong-Soon Kim*, Jae-Gyun Gwag, Yu-Mi Choi, Ki-Hoon

Park and Chang-Ki Shim. Agrobiodiversity Center, Rural Development of Administration, Swon 441-853, Korea

About 2,760 accessions of four species of Tomato germplasm were conserved in Agrobidiversity Center of RDA, Korea. Phytophthora root rot caused by the soil born pathogen, Phytophthora capsici, is one of the most important diseases of Solanaceae crops including tomato. We investigated the resistant of 200 accessions of tomato germplasm against P. capsici. About 70 days old seedlings were used and replicated 15 plants each accession. The inoculums concentration was adjusted to 10 zoosporangium/ml by haemocytometer. The disease incidence was checked from 7days to 28days at an interval of 7days after inoculation. Among 200 accessions, incidence of phytophthora blight of the 94 accessions of tomato (ca. 47%) was more than 40%, while no disease was observed in 64 accessions (ca. 32%) at 7days after inoculation. At 28 days after inoculation, the 48 accessions of tomato do not infected with P. capsici, while the 77 susceptible accessions of tomato were scored with more than 70% incidence of phytophthora blight. In the susceptible accessions of tomato, abundant sporangia were formed plant stem, while no sporangia were produced on the stem and root which were found resistant. This result suggests that the 48 accessions of tomato germplasm might be used as 5

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the breeding resources for the candidate of phytophthora resistance breeding grogram. Further genetic study should be carried out to verify this result. And this study was established with the overall focus of providing information on important characters of the genetic germplasms.

G-20

Evaluation of Powdery Mildews resistance caused by Erysiphe spp in Squash germplasm (Cucurbita spp.). Jeong-Soon Kim , 1

Woo-Moon Lee , Jae-Gyun Gwag , Ki-Hoon Park and Chang-Ki Shim . National Agrobiodiversity Center, Rural Development of 2

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Administration, Suwon 441-853, Korea; National Institute of Horticultural & Herbal Science, Rural Development of Administration, Suwon 475, Korea 2

Above 2,500 accessions of Squash germplasms, Cucurbita maxima, C. maxima subsp. maxima, Cucurbita mixta, Cucurbita moschata, Cucurbita pepo, and C. pepo subsp. pepo, is conserved in Agrobidi-

versity Center of RDA, Korea. We investigated the powdery mildew resistance of the 117 accessions of 3 species of Cucurbita spp. germplasm, C. maxima, C. moschata, and C. pepo, were estimated with disease incidence. The disease incidence was scored as percentage of the below 10 leaf area infected at 15 days intervals from July 15, 2009 to September 14, 2009. The symptom of disease appeared spontaneously at July 21, 2009 without chemical control. Among 117 accessions of squash germplasm, incidence of powdery mildew of the 105 accessions of tomato (85.4%) was more than 32% of leaf area infected, while no disease was observed in 12 accessions (10.2%) at 54days after disease occurrence. In the susceptible accessions of squash, abundant conidia were formed plant leaf and stem, while no symptom and conidia were produced on the plant which was found resistant accessions. The 100% of disease incidence was recorded in 52 accessions (44.4%) of C. pepo and each of 16 accessions (13.7%) of C. maxima and C. moschata, with affected leaves accounting for more than 80% of leaf area infected with Erysiphe spp. Resistant to the disease without symptom were found 9 accessions of C. maxima, 1 accession of C. moschata, and 2 accessions of C. pepo. Further PCR identification and detection should be carried out to verify this result. And this study was established with the focus of providing information of the squash germplasm for the breeding of powdery mildew resistance new varities.

G-21

Investigation of Phytophthora Blight caused by Phytophthora capsici for evaluation of Hot Pepper germplasm (Capsicum annum). Jeong-Soon Kim*, Jae-Gyun Gwag, Ki-Hoon Park and Chang-Ki Shim. Agrobiodiversity Center, Rural Development of Administration, Suwon 441-853, Korea

Phytophthora Blight caused by Phytophthora capsici, is one of the most destructive diseases to the yield decrease of Solanaceae crops. About 4,405 accessions of Pepper germplasm were conserved in Agrobidiversity Center of RDA, Korea. We investigated the resistant of 300 accessions of hot pepper germplasm against P. capsici. About 70 days old pepper seedlings were investigated with 18 replicate each accessions, and inoculated with the concentrations of 10 zoosporangium/ml adjusted by haemocytometer. Five resistant varieties of 5

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commercial pepper, Yeogkanghongjanggun, Buja, Umchungna, Powerspeed, and PR-Daechon, and two susceptible varieties of commercial pepper, Hongboseg, and Gumdang, were used as control plants. The disease incidence was checked from 7days to 28days at an interval of 7days after inoculation in the greenhouse conditions. Among 300 accessions, incidence of phytophthora blight of the 67 accessions of pepper (22.3%) was more than 60.1%, while no disease was observed in 37 accessions (12.3 %) at 7days after inoculation. At 28 days after inoculation, the 5 accessions of pepper do not infected with P. capsici, while the 244 susceptible accessions (81.3%) of pepper were scored with more than 60.1% incidence of phytophthora blight. In this study, five resistant germplasm against P. capsici is a domestic landrace pepper accessions. This result suggests that the 5 candidate pepper germplasm might be used as the breeding resources for phytophthora resistance breeding program. Further genetic study should be carried out to verify this result and was established with the overall focus of providing information on important characters of the pepper germplasms.

G-22

Selection of Field Resistance Variety to Bacterial Leaf Blight of Rice. Du Ku Lee, Tae Hwan Noh, Mi Hyung Kang, Chae Hoon Paik, Geon Hwi Lee, Si Ju Kim and Jae Duk Kim. National Institute of

flower all over the world. It's economic influence and capability was very big. So we collect 18 carnation varieties which mainly circulated in the market place and set the best way for creating more efficient tissue culture system to get much more plant organism through micro propagation. 4 Carnation(Dianthus caryophyllus) varieties Yellowdotcom, Jakarta, Belmonte, Polartessino, were selected for having more organ division capacities from one cell line. The common fittest complex growth regulator combinations through 4 varieties was MS+Sucrose 3% + NAA 1.0mg/L+TDZ 1.0mg/L. and S+Sucrose 3%+NAA 3.0mg/L+BA 1.0mg/L for Belmonte. To get a most efficient callus induction and regeneration capacity we have to use MS as a basal culture medium and Sucrose 3% as a carbon source. The coagulating agent for get more plantlets from callus was Phytagel and concentration was 0.3%. The best culture material for Carnation (Dianthus caryophyllus) was stem. when we used this part as culture material we can elevated shoot formation rate by 80.2% contrast to leaf culture and culture periods was shortened by 6 days. Based on this result we insert 10 RNAi gene to bring up the genetically modified Carnation(Dianthus caryophyllus) for being intensified resistive to root-knot nematode and get selection marker resistance 346 Carnation(Dianthus caryophyllus) transgenic lines from 4 varieties ready for bioassay.

Crop Science, RDA, Iksan 570-080, Korea

G-24

We selected field resistance varieties to baterial leaf blight (BLB) of rice caused by Xanthomonas oryzae pv. oryza. The field resistance varieties of BLB were Samkwang, Hopum, Haechanmulkyeol, Saenuri, Hwangkeumnuri and Hwangkeumnodul. For selection field resistance varieties, three tests were used to carry out vertical resistance, secondary infection and secondary transmission test. The result of vertical resistance test were selected Samkwang, onnuri, Gangbaek, Jinbaek, Unmi, Hopum, Heachanmulkyeol, Borimi, Saenuri, Hwangkeumnuri and Hwangkeumnodul, that showed lower than 1.5cm of disease lesions length. Samkwang, Hopum, Gangbaek, Jinbaek, Saenuri, Hwangkeumnuri and Hwangkeumnodul were lower than 2.5% secondary infection rate. In secondary transmission test, Saenuri, Jinbaek and Hwangkeumnodul were not transmitted; however, Unmi, Hopum, Borami, Hwangkeumnuri and Heachanmulkyeol were lower than 6.2% transmitted respectively.

Kyoung Han, Jong-Han Park, Kyung-Sook Han, Dae-Hyun Kim, Jung-sup Lee and Jeong-Hwan Hwang. Horticultural & Herbal Crop

G-23

The selection of Optimum Carnation(Dianthus caryophyllus) Varieties and Setting The culture condition for Bringing up the Genetically Modified Crops for Being Intensified Resistive to Root-knot nematode. Chan-Ho Kang *, Soo-Gon Han, Sung-Hwan

Resistant Cultivar Screening to Bacterial wilt of tomato. YouEnvironment Division, National Institute of Horticultural & Herbal Science, RDA, Suwon 441-440, Korea

Bacterial wilts, caused by Ralstonia solanacearum, is a very damaging disease to tomato plants (Lycopersicon esculentum Mill.) in Korea. Selection of resistant cultivar is the best way to prevent or reduce the occurrence of the disease. It could be used to control Bacterial wilts in tomato if resistant cultivars could be identified or developed. Twenty-two cherry tomato cultivars, thirty nine tomato cultivars and thirteen parent stem tomato cultivars were inoculated with Ralstonia solanacearum, to evaluate tomato cultivar-specific resistance. Thirty six cultivars were highly susceptible to bacterial wilts; 61-100% of their whole tissue became wilted within 7 days after inoculation. Twenty one cultivars and sixteen cultivars were the moderate resistant and resistance to bacterial wilts. In an evaluation of 73 major commercial cultivars, ‘Lilyance’, ‘TP-7’, ‘Choice’, ‘B-blocking’ and ‘Friends’ were found to have a high level of resistance to bacterial wilts of tomatoes.

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Kwon, Young-Ju Song and Bum-So Han . Jellabukdo Agricultural Research & Extension Service, Iksan 570-080, Korea; National Academy of Agricultural Science, Rural Development of Administration, Suwon 441-853, Korea 2

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G-25

Re-classification of Soybean Differentials using 11 strains of Soybean Mosaic Virus. Yeong-Hoon Lee *, In-Youl Baek , Sang1

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Through molecular breeding progress, many plant transformation techniques for improving transformation accuracy to get more useful transgenic plants were activated. But hour by hour, new kinds of plant and varieties for new needs were developed continuously. so the new transformation techniques faced with newly developed varieties were also requested. Carnation(Dianthus caryophyllus) was main ornamental

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Ouk Shin , Byong-Won Lee , Hyun-Tea Kim , Jong-Min Ko , SeaGyu Lim , Ha-Sik Shim , Myeong-Gi Jeon , Won-Young Han , Young-Nam Yoon , Jung-Kyung Moon , Su-Heon Lee , Gey-Woon Lee and Keum-Yong Park . Department of Functional Crop, NICS, 1

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RDA, 1085 Neidong, Miryang, 627-803, Korea; Plant Pathology Division, NIAST, RDA, Suwon, 441-707, Korea; Department of Agricultural Biology, Kyungpook National University, Daegu 702701, Korea 2

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Abstract of Presentations Soybean mosaic virus (SMV) is a prevalent pathogen that causes significant yield reduction in soybean [Glycine max (L.) Merr.] produc-

tion worldwide. SMV belongs to potyvirus and causes symptoms such as mild mosaic, mosaic and lethal necrosis. SMV is seed-borne and is also transmitted by aphids. They classified 98 SMV isolates from the USA soybean germplasm collections into strain groups, G1 to G7, based on the symptoms induced on a set of soybean differential cultivar. In Japan, 102 SMV isolates were classified into 5 strains, A to E, based on differential symptoms induced on 4 soybean cultivars. In China, 8 SMV strains, S through S , in the Mid-south region were isolated and characterized. Lu et al. (1985) also reported three SMV strains (SMV-1 to 3) in the Northeastern China. We have used the system of Cho and Goodman's differential reaction for seven strains of SMV. After the initial grouping of SMV, several resistance-breaking isolates have been identified during the last two decades in Korea. In other to re-classification of SMV differentials, we inoculated 11 strains (G1-7, G5H, G6H, G7H and G7a) on fifty-four cultivars. These cultivars were classified into 18 patterns through pathogenic response such as resistance, necrosis and susceptibility. In addition, 8 resistant and 6 susceptible cultivars were confirmed against all strains of SMV. a

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G-26

Resistance to Fusarium head blight caused by Fusarium graminearum in Synthetic wheat. Hyunkyu Sang , Kwanggeun 1,2,3

Park , Sangmi Yu , Etienne Duveiller and Roman Delgado . 1,3

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International Maize and Wheat Improvement Center, Apdo.Postal 6641, 06600 Mexico, D.F., Mexico; College of Agricultural and Life Sciences, Chungnam National University, Daejeon 305-764, Korea; Rural Development Administration, Suwon 441-707, Korea 1

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Agricultural Microbiology Division, National Academy of Agricultural Science, RDA, Suwon 441-707, Korea; Department of Agricultural Biotechnology, Center for Fungal Genetic Resources, Seoul National University, Korea 1

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Rice blast caused by Magnaporthe oryzae, is the most widespread and threatening disease for rice, which causes significant crop losses annually throughout the world. To investigate avirulence (AVR) genes, random insertional mutants of M. oryzae strain KJ201 (~16,000 isolates), were generated via Agrobacterium tumefaciensmediated transformation (ATMT) in the previous study (Jeon et al., 2007) and they were used for a bulk inoculation test. Seventeen mutants were finally selected through morphological, physiological and pathogenicity tests. Pathogenicity of the selected mutants was evaluated using 31 monogenic rice lines, which lines confer single major blast resistant gene. Mutants with loss of virulence mutants had higher numbers than those with gain of virulence (GOV). Mutant lines, GOV30, GOV32, GOV37, GOV40, GOV41, and GOV50, were increased their virulence to major rice resistance genes including Pik-s, Pik, Pik-p, Pib, Pi7, Pi11, and Pita-2. Knock out mutants for the corresponding resistant genes are under construction to validate whether the disrupted genes are avirulence genes or not.

G-28

Studies on QTL-Associated Genes to Improve Multigenic Disease Resistance in Rice. Se-Weon Lee, Jae-Yun Jeong, Zu-An Lee, Yong

Hwan Lee, Hong-Sik Shim, Dong-Soo Ra, and Seong-Sook Han. Agricultural Microbiology Division, National Academy of Agricultural Science, RDA, Suwon 441-707, Korea

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Fusarium head blight caused by Fusarium species, including Fusarium graminearum, F. culmorum, F. avenaceum, F. poae and Microdochium nivale, is one of the most destructive diseases of wheat in

regions with wet conditions. The disease may cause serve yield reduction and can occur high levels of mycotoxin contamination such as deoxynivalenol, causing mycotoxicosis in human and domestic animals. Although several species of Fusarium are capable of inciting FHB in wheat, F. graminearum is the most dominant species in Mexico. We present the results of resistance to FHB caused by Fusarium graminearum in Synthetic wheat. Synthetic hexaploid wheat lines were made at CIMMYT between Aegilops tauschii accessions from several geographical regions and various durum wheats from the CIMMYT program. Resistance testing was performed in a field trial artificially inoculated with a F. graminearum conidial suspension. FHB index was assessed 30 days after inoculation on 5 spikes per plot tagged at anthesis. The results revealed that 23.1% of SHW lines were less than 5% of the score for FHB index. These SHW lines demonstrated a great potential to enhance resistance of wheat to FHB. Also 57.4% of SHW lines were 5–30% of the score for FHB index and 19.5% of SHW lines were more than 30% of the score for FBH index.

G-27

Characterization of Gain of Virulence Mutants to Find Avirulence Genes in Magnaporthe oryzae. Zu-An Lee , Myoung-Hwan 1

Chi , Jae-Yun Jeong , Yong Hwan Lee and Se-Weon Lee . 2

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Rice blast and bacterial blight, caused by Magnaporthe oryzae (Mo) and Xanthomonas oryzae pv. oryzae (Xoo), respectively, are the most destructive fungal and bacterial diseases of rice worldwide. Genetic resistance is the most effective way to control the diseases. However major resistance is often broken down within a few years after release of new cultivars due to the high degree of pathogenic variation. In this study, we analyzed gene expression patterns using microarray and predicted the location of the highly expressed genes on chromosomes to find QTL-associated genes. Palgongbyeo, known as a durable resistant cultivar, was selected for the rice blast resistance study. Gene expression profiles of the Palgongbyeo inoculated with 19 mixed Mo isolates was analyzed using microarray. Of 27,614 genes on the chip, over 2,700 genes were up-regulated after the inoculation. We selected 199 genes that constitutively up-regulated at least two fold all detection time points. The 199 up-regulated genes were involved in various functions such as information storage, cellular processing, and metabolism. Especially, many genes were related in secondary metabolites biosynthesis transport and signal transduction mechanisms. The selected 199 genes were mapped on an existing linkage map consisting of rice blast QTL. The genes are well distributed on the twelve rice chromosomes and 38 genes are associated with blast resistant QTL. Microarray analysis for bacterial blight resistance is underway to find useful candidate genes conferring both Mo and Xoo broad spectrum resistance.

G-29

Recent Race Distribution Shift of Magnaporthe oryzae in Korea.

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Dong-Soo Ra, Se-Weon Lee, Yong Hwan Lee, Hong-Sik Shim and Inn-Shik Myung. Agricultural Microbiology Division, National Academy of Agricultural Science, RDA, Suwon 441-707, Korea

To investigate the distribution and frequency of blast pathogen races, diseased leaf and panicle samples were collected from farmer' field in Korea. Using Korean differential varieties, 973, 853, 1,016 and 705 isolates from 2005 to 2008, respectively, were differentiated. Nationwide frequency of KJ races was 75.6%, 81.7%, 84.4%, and 81.8% from 2005 to 2008, respectively. KJ race frequency of Gyeonggi province was 85.7% (2005), 69.0% (2006), 76.5% (2007), and 81.8% (2008); and was 71.5% (2005), 98.6% (2006), 99.2% (2007), and 95.1% (2008) in Gangwon province. KJ race frequency of Chungbuk province was 94.9% (2005), 81.36% (2006), 90.2% (2007), and 88.0% (2008); and was 64.7% (2005), 85.3% (2006), 69.8% (2007), and 67.0% (2008) in Chungnam province. KJ race frequency of Jeonbuk province was 82.4% (2005), 93.1% (2006), 75.5% (2007), and 70.5% (2008); and was 87.9% (2005), 97.5% (2006), 81.1% (2007), and 90.1% (2008) in Jeonnam province. KJ race frequency of Gyeongbuk province was 61.0% (2005), 59.0% (2006), 94.4% (2007), and 81.2% (2008); and was 61.2% (2005), 89.9% (2006), 87.9% (2007), and 82.7% (2008) in Gyeongnam province. In general, KI race frequency were lower than KJ race, but some local regions such as Gyeonggi, Chungnam, Gyeongbuk, and Gyeongnam provinces showed over 30% of KI race frequency. The most frequently isolated races were 101, 201, 105, and 301 of KJ and 1113, 1117, 409, and 413 of KI, but race distribution patterns varied a lot between each year.

G-30

Accumulation of scoparone, a phytoalexin associated with resistance to plant pathogens in citrus leaves inoculated with citrus scab pathogen(Elsinoe fawcettii). Pyoungho Lee, Jae-wook Hyun,

Jihyun Lee, Hyojung Kim and Young-Jin Kim. Citrus Research Station, National Institute of Horticultural & Herbal Science, RDA; Division of Life Science, College of Life Science & Biotechnology, Korea University, Seoul, Korea, 136-701 1

It has been known scoparone is a phytoalexin associated with the resistance of citrus against plant pathogenic fungi. In inoculation test in greenhouse condition, the symptoms were firstly appeared 3 days after inoculation with citrus scab pathogen, Elsinoe fawcettii on leaves of one-quarter mature size of satsuma mandarin(Citrus unshiu Marc.). The scoparone accumulation was detected 24 hr after inoculation but a little was accumulated 12, 36, 48, and 76hr after inoculation in first experiment, and it was 36 hr in second experiment. And the scoparone was also detected on other noninoculated flush by inoculation on leaves of one branch of same tree. This may means that the citrus leaves have SAR against scab pathogen.

G-31

The non-protein amino acid B-aminobutyric acid (BABA), known as an inducer of resistance against infection of plants by various pathogens and abiotic stresses. We have isolated gene encoding non-protein amino acid B-aminobutyric acid (Brbaba) from Brassica rapa, characterized with gene expression pattern analysis. Also we expressed (Brbaba) gene in the tobacco through Agrobacterium tumefaciens mediated transformation methods using pBig binary vector system which has modified cauliflower mosaic virus (CaMV) 35S promoter. The transgenic plants was confirmed by PCR analysis, Southern blot, the gene expression in transgenic plants was investigated by RT-PCR analysis respectively. We are investigating the effect of (Brbaba)induced protection against fungus and abiotic stresses tolerance in transgenic tobacco plants. The preliminary results suggest that transgenic tobacco plant which expressed B-aminobutyric acid (Brbaba), showed enhanced resistance to fungus pathogens and abiotic stresses.

H-1

Disease models for timing fungicide applications to control anthracnose crown rot of strawberry. Myeong-Hyeon Nam , N. A. 1

Peres , S. J. Mackenzie , In-ha Lee and Hong-Gi Kim . Nonsan Strawberry Experiment Station, Chungnam ARES, Nonsan, 320-862, Korea; Department of Gulf Coast Research and Education Center, University of Florida, Wimauma, FL, 33598 USA; Department of Applied Biology, Chungnam National University, Daejeon 305-764, Korea 2

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Anthracnose crown rot (ACR) of strawberry caused by Colletotrichum gloeosporioides is a serious problem in Korea and in the southeastern United States, because these regions are relatively warm and have sufficient rainfall to spread the disease. Regression analysis of temperature (T) and wetness duration (W) on the proportion of strawberry plants infected (Y) was determined in growth chamber studies with cv. Seolhyang. Seven-day captan spray schedule, logistic infection model + early detection by real-time PCR, TOM-CAST weather forecasting system, and infection risk model using temperature and rainfall are being evaluated in nursery field. The logistic regression equation for C. gloeosporioides was In(Y/[1 - Y]) = -1.82 + (0.0028WT). Most of the model-based spray programs that included applications prior to symptom development reduced disease incidences below 2% compared to 10% observed in the control plots. The best treatment was the one in which logistic infection model + early detection spray program by real-time PCR was used. This treatment reduced the number of fungicide applications by 50% or more compared to the current calendar-based application. The development of real-time PCR-based early detection conjugated with logistic infection model will enable growers to have the information necessary for them to decide on the need for a fungicide application.

H-2

A forecasting system for fruit disease and insect pest occurrence on the real-time basis. Yong kyu Han , Yong Soon Shin , Wee Soo 1

Isolation and functional analysis of a gene (Brbaba) encoding Non protein amino acid Beta-aminobutyric acid from Brassica rapa L. Uuganchimeg bayarsaikhan , Yu Jin Jung , Bayarbat 1

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Ishvaanjil , Inhye Lee and Kwon Kyoo Kang . The Graduate School 3

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of Biotechnology and Information Technology, Hankyong National University, Ansung City 456-749, Korea

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Kang , Kye Chan Choi , Jong Haeng Heo , Hong Deok Chae , Eok Geun Jeong , Soon Sung Hong , Sung Ki Kim and Eun Woo Park . 2

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EPINET Co., Business Incubator, Seoul National University, Suwon, 441-853, Korea; Department of Agricultural Biotechnology, Seoul National University, Seoul, 151-921, Korea; Chungcheongnam-do Agriculture Research and Extension Services, Yesan, 340-861, Korea; 1

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Abstract of Presentations Cheonan-si Agricultural Technology Center, Cheonan, 330-846, Korea; Gyeonggi-do Agricultural Research and Extension Services, Hwasung, 445-972 4

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A forecasting system (http://www.cnipm.kr) was developed to provide information on possible outbreaks of diseases and insect pests for fruit growers at seven locations in Chungnam Province. This system, which consists of weather data acquisition system, data storage system, job process system, web service system, and short message service (SMS) system, generates and distributes information through Internet on hourly weather conditions, and disease and insect pest forecasting on the real-time basis. It also provides short warning messages on disease and insect pest forecasting to cellular phones of fruit growers and other end-users. The hourly weather conditions are monitored by automated weather stations installed at fruit tree orchards. The weather data include hourly temperature ( C), rainfall (mm) and leaf wetness (fraction of an hour). Disease and insect pest forecasting information are available for five apple diseases (white rot, bitter rot, Marssonina blotch, scab and Alternaria blotch), two pear diseases (scab and rust), nine insect pests damaging apples and pears (comstock mealybug, psylla, oriental fruit moth, leaf miner, peach fruit moth, asiatic leafroller, leaf-curling aphid, two-spotted spider mite, and European red mite), and two diseases of grapevines (bitter rot and downy mildew). The system is programmed to send short message warnings to users cellular phones at 09:00 am everyday if possible occurrences of diseases and insect pests are forecasted. o

H-3

Strategies for managing Bakanae disease epidemics of rice through the basic reproduction number of the pathogen. K. Ishiguro , T. Hayasaka , K. Kurauchi , N. Hatanaka and H. Horaguchi . National Agricultural Research Center, Tsukuba 3051

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8666, Japan; Yamagata General Agricultural Research Center, Tsuruoka 999-7601, Japan; Aomori Prefectural Industrial Technology Resarch Center, Kuroishi 036-0522, Japan; Miyagi Prefecture Furukawa Agricultural Experiment Station, Osaki 989-6227, Japan; Iwate Agricultural Research Center, Kitakami 024-0003, Japan 2

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In Tohoku region in Japan, epidemics of Bakanae disease of rice, a seed-borne disease caused by Gibberella fujikuroi, have recently become prevalent. Adding to the seed-borne nature, spores of the pathogen formed on the diseased stems of rice in the fields disperse as air-borne spores to rice panicles, which cause infected seed. Some of certified seed, which majority of the growers use, may be contaminated by some infected seed, while regulation to help prevent the contamination has been strictly applied in the certified seed production program. Consequently, spores dispersed from diseased rice plants in the nearby grower fields to those for certified seed production should be the probable infection sources of the certified seed. Recently, hotwater treatment for seed disinfestations and/or biological control agents for seed treatment become widespread as alternative measures for chemical seed treatment, while inaccurate operation often causes not enough outcomes. To decline the epidemics effectively and efficiently, we roughly estimated the basic reproduction number (R ) of the pathogen using several tentative values, and tried to determine which factors to significantly reduce R are. The pathosystem includes a disease chain between certified seed and rice on grower fields, and another chain within rice on grower fields via self-produc0

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ing seed. Because the disease cycle is complex as such, we set up two equations and then converted them into a matrix population model. The dominant eigenvalue of the matrix was considered to be R . Tentatively, for example, low temperature storage of seed after hot-water treatment and increase proportion of certified seed use seems to be effective to reduce R . Further experiments to quantify multiplication factors of each process in the pathosystem will be needed to estimate more accurate and precise R , and hence to explore more realistic management strategies of the disease. 0

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H-4

Potential risks of exotic forest fungal pathogen introduced by imported timbers. Nayoon Huh, Min-Ji Kim, Gyu-Hyeok Kim and Jae-Jin Kim. Division of Environmental Science & Ecological Engineering, Korea University, Seoul 136-713, Korea

As the global demand for forest products increases, importation and exportation of logs throughout the world have been increasing rapidly. The increase of the imported logs from other country can bring introduction of additional pathogens from foreign pests for which they have no natural protection. When these foreign pests bring new diseases into the country, they can destroy not only wood products or plants, but the whole ecosystems. Therefore, in our study, fungal speciess were isolated from Picea abies, Picea jezoensis and Pinus radiata imported from north to eastern Siberia, Europe and New Zealand, respectively. Also we established their biodiversity and investigated the present status and/or potential risk as tree pathogen of those identified fungal species. A total of 342 fungal isolates were obtained and identified using morphological and molecular biological methods. As a result, 53 species of ascomycetes and 9 species of basidiomycetes including 8 unknown species were identified. Among them, at least 9 species including Hormonema dematioides, Leptographium proceum, Nectria sp. Ophiostoma piceaperdum, Ophiostoma nigrocarpum, Ophiostoma pluriannulatum-like, Valsa kunzei, Lenzites betulina and Tyromyces chioneus were considered to be warning pathogens further in other countries as Australia, because of their serious risk potential causing wood discoloration or wood rot of trees and wood products, either plant diseases of aboveground tree part. However, they have not been introduced as tree pathogens in Korea yet, hence they are being brought into Korea without any boundaries or restrictions.

H-5

The control efficacy of pear rust based on real time forecasting model of internet hompage. Soon Sung Hong, Jin Young Kim, Jin Gu Lee, Hyun Ju Lee and Sung Kee Kim. Gyeonggi-do Agricultural Research and Extension Service, Hwaseong 445-784, Korea

The real time pear disease forecasting system was established in the internet homepage “www.epilove.com” based on weather data collected from automatic weather station(AWS). The timing of teliospore dispersion on Juniperus chinensis was predicted from forecasting model by average temperature of March. The probability of pear rust was divided 4 grades depending on rainfall and raining time before and after the timing of teliospore dispersion. The model forecasted the date of occurrence of pear rust, May 1, 2007 in the southern part of Gyeonggi, April 23, 2008 in the east and north part of Gyeo-

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nggi and April 20 to 25 in 2009 in the middle and south part of Gyeonggi. The control efficacy of pear rust was from 89.9 to 100% by spraying the Hexaconazole 2% based on forecasting model on May 2 in 2007, April 23 in 2008 and April 21 in 2009. This system will be very helpful to reduce the fungicide by controlling pear rust on optimum time predicted from pear disease forecasting model.

H-6

Diversity of Airborne Fungi at Oriental Pear Farms and Their Growth Characteristics. Hye Yeon Mun and Hyang Burm Lee. Division of Applied Bioscience & Biotechnology, College of Agriculture & life Sciences, Chonnam National University, Gwangju 500757, Korea

This study was performed to investigate the seasonal diversity of airborne fungi at oriental pear [Pyrus pyrifolia (Burman) Nakai] farms located in Naju, Jeonnam Province, Korea. During February 2007 to February 2008, the outdoor samples of airborne fungi were taken in two locations of the pear farms using a plate lid opening method for 10 to 30 min (up to 3hr) and then incubated at 23 C for 7-14 days. Each isolate was identified based on cultural, morphological characteristics and 18S and 28S ribosomal RNA gene analyses. Alternaria, Cladosporium, Epicoccum, Fusarium and Periconia were most prevalent airborne fungi. Aspergillus and Penicillium are the next one. Some coelomycete fungi including Phoma and Phomopsis, and other genera such as Leptosphaeria, Phaeosphaeria and Phaeosphaeriopsis belonging to order Pleosporales were also frequently detected in the air samples. Various growth characteristics of the airborne fungi were investigated and will be shown in this poster. o

H-7

Diversity of Endophytic Fungi on Juniperus chinensis in Korea. Hyung Ryul Jang and Hyang Burm Lee. Division of Applied Bio-

science & Biotechnology, College of Agriculture & Life Sciences, Chonnam National University, Gwangju 500-757, Korea

The diversity of fungal endophytes associated with leaf and twig of Chinese juniper (Juniperus chinensis L.) on campus of Chonnam National University, Gwangju, Korea was investigated during July 2007 - May 2008. In total, 160 isolates were isolated from the twigs and leaves of the plant. The isolates were identified by morphological characteristics and 18S and 28S rRNA gene analyses. Out of 160 endophytic fungi, 43 isolates belonged to Pestalotiopsis species which was dominant endophyte. Fungal diversity in relation to culture morphology as well as to host and season association will be discussed.

H-8

Differences between two fungal species, Collectotrichum gloeosporioides and C. acutatum causing apple bitter rot, in the sensitivities to fungicides using for apple in Korea. Kyung Jin Jun, Gun Woong Kim and Jae Youl Uhm. School of Applied Biology and Chemistry, College of Agricultural and Life Sciences, Kyungpook National University, Daegu, 702-701, Korea

The differences in the sensitivities between two fungal species caus-

ing apple bitter rot, Collectotrichum gloeosporioides and C. acutatum, to the fungicides using in the current spraying program for apple in Korea were examined. Fungal isolates collected in 2002 and 3 successive years from 2006 were subjected to species classification on the basis of sensitivities to benomyl and diethofencarb. The distribution frequency of C. acutatum in 2002, until which benzimidazole fungicides had intensively used in most of apple orchards showed a wide variation by sites, ranging from 0 to 92.3%, average 42.2%. Sensitivities of the fungicides to both species were assessed by EC values for mycelial growth. EBI and QoI showed higher sensitivity to C. acutatum than C. gloeosporioides but iminoctadine-triacetate and fluazinam were the reverse, especially iminoctadine-triacetate was almost totally insensitive to C. acutatum. EC values of kresoxim-methyl and trifloxystrobin for C. gloeosporioides were widely distributed but those of pyraclostrobin were not. Isolates in C. acutatum of which mycelial growth were not inhibited at high concentration of dithianon were sporadically appeared at first, but their frequency was gradually increased. Similar phenomenon was observed in some isolates of C. gloeosporioides for iminoctadine-triacetate of which EC value for mycelial growth were largely shifted from average were began to appear from 2007 and become more conspicuous in 2008. As those isolates of which mycelial growth was not inhibited at high concentration are increasing, the effects of them on the control of bitter rot are required to be monitored. 50

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H-9

Seasonal Occurrence of Peach Diseases Past 10 years. Jong-Han

Park, Kyung-Sook Han, You-Kyoung Han, Jung-Sup Lee, Dae-Hyun Kim and Jeong-Hwan Hwang. National Institute of Horticultural & Herbal Science, RDA, Suwon 441-440, Korea

Seasonal occurrence of diseases of peach trees in orchards was investigated from 1999 through 2008 at three locations, Suwon, Eumseong, Icheon, and Yeongi in Korea. The examination was carried out two times per year for ten years. As a result of the survey, the major diseases of peacjes were bacterial shot hale caused by Xanthomonas campestris pv. pruni and leaf curl caused by Taphrina deformans. Between the data obtained from this study and weather condition were analysed. There were high relation between diseases occurrence and times of the rainfall per month. The other diseases occurring peach orchards were anthracnose caused by Colletotrichum gloeosporioides, brown rot caused by Monillonia fructicola, die-back caused by Botryospheria dothidea or Phomopsis sp., crown gall caused by Agrobacterium tumefaciencs, and powdery mildew caused by Sphaerotheca pannosa et cetera. The result of this study can be provide basal and important information on the setting-up diseases management program for the purpose of Good Agricultural Practice (GAP).

H-10

A Forecasting Model for Estimating the Initial Date of the Late Blight Occurrence in Pepper Fields after Overwintering Periods. K. S. Do , S. C. Yun and E. W. Park . Dept. of Agricultural Biotech1

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nology, Seoul National University, Seoul, 151-921, Korea; Dept. of Biomedical Sciences, Sun Moon University, Asan, Chungnam, 336708, Korea 2

Abstract of Presentations A forecasting model for estimating the initial date of the late blight occurrence in pepper fields was developed based on the assumption that the germinated zoosporangia or zoospores can survive three days in soil, one zoosporangium produces 20 zoospores and the direct germination of the zoosporangium is only affected by soil temperature. The disease risk level (RL) to estimate the initial date depends on the 3-day moving average of the amount of active inoculum in soil (AI). The AI was calculated as AI=(1+19p)·Z, where p is the ratio of the zoosporangia which release zoospores, and Z is the amount of zoosporangia remained in soil at a day. In the equation, p is a temperature function, and Z depends on both temperature and soil water content. The volumetric soil water content was estimated from air temperature, relative humidity, rainfall, soil texture, and the crop basic coefficient by using the SCS runoff model and the David evapotranspiration model. Based on the analysis on 18 sets of weather and disease survey data obtained from 14 locations in 2001 to 2003, we were able to determine that the disease could occur at 5 days after RL became greater than or equal to 68 after overwintering. When the disease forecasting model was evaluated using the weather and disease survey data obtained from 14 locations in 2004 to 2009, the predicted dates of first occurrence of the disease after overwintering were within the range of 10 days from the observed dates except for one.

H-11

Outbreak of Bakanae disease of rice and its cultural and meteorological factors in Korea. Joo Hong Lee, Yong Hwan Lee, Woo-

Sik Park, Se-Weon Lee and Hong-Sik Shim. Agricultural Microbiology Division, National Academy of Agricultural Science, Rural Development Administration, Suwon, 441-707, Korea

This study was conducted to analyze the cultural and meteorological factors on affecting the sudden increasing of bakanae disease of rice in Korea. Bakanae disease occurred a little in early maturing varieties of rice from late of 1990s but it was not considered main disease by farmers because of very low cultivation areas (below 1%) of early maturing varieties of rice. In 2004, this disease was suddenly showed on nursery boxes all over the nation and then 12.1% of severely infected nursery boxes, seeded ‘Junambyeo’ variety produced in Asan, Chungnam provinces, appeared in Sangju, Kyeongbuk provinces in 2005. Spores of bakanae disease dispersed on the surface of bottom of infected tillers and then infected to rice spikelet during heading period. Infection of spores to rice spikelet was severe in high average temperature over 25 C during heading period. Average temperature from late July to middle August was generally around 25 C and then down below 25 C at late August in Korea. And middle late maturing varieties of rice, which heading period was late August, were cultivated over 75% in Korea. It was tendency that transplanting time was earlier and then rice heading time shifted to middle of August in southern and western coast part of Korea after 2000s. In 2003, especially, average temperature was higher 2-4 C than normal year in middle of August. It was thought that the high temperature during rice heading period by the early transplanting of rice was one of the factors of outbreak of bakanae disease in Korea. o

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H-12

Review of incidence of rice major diseases in Korea from 1999 to

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2008. Yong Hwan Lee*, Dong-Su Ra, Hong-Sik Shim, Wan-Hae

Yeh, Hyo-Won Choi, Sung-Kee Hong, Young-Kee Lee, Inn-Shik Myung, Se-Weon Lee, Sang-Yeop Lee, Jin-Woo Park, Hong-Su Choi, Su-Heon Lee, Wan-Gyu Kim, Dong-Su Ra, Hong-Sik Shim, Kyung-Seok Park, Jeong-Soo Kim, Yong-Gi Kim and Sung-Suk Han. Agricultural Microbiology Division, National Academy of Agricultural Science, Rural Development Administration, Suwon, 441707, Korea

In the period of 1999 to 2008, we surveyed the diseases incidence of rice in Korea. Leaf blast was shown 0.85% of the diseased leaf area in 2000 and then consistently decreased 0.02% in 2008. It was thought that amount of applied nitrogen fertilizer was dramatically decreased from 18 kg/10a to 11kg/10a by policy of local governments after 2000. But, panicle blast was irregularly appeared by the years as the weather condition during heading to milk stage of rice. Diseased lesion height of sheath blight was the lowest (7.7%) in 2003 and the highest (21.4%) in 2007. It was shown high incidence in condition of high temperature and sunshine from middle of August to early of September. Brown spot occurrences in Chunbuk, Gyeongbuk, and Gyeongnam provinces had been on the decline since 2001, but those in Chungnam, Jeonbuk, and Jeonnam provinces had increased since 2005. Falls smut was drastically decreased after shown 7.59% of diseased panicles in 2000. The correlation coefficient between diseased panicles of false smut and rainfall during last August was 0.77*. Bacterial blight was severely increased at the regular diseased field areas in Jeonnam, Jeonbuk, Gyeongnam, and Chungnam province since 2003.

I-1

Studies on molecular plant-microbe interactions in Japan Aspects on phytopathogenic bacteria. Shinji Tsuyumu and Hisae 1,2

Hirata . Graduate School of Science and Technology, Shizuoka Uni2

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versity, 836 Ohya, Suruga-ku, Shizuoka 422-8529, Japan; Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka 422-8529, Japan 2

For the complex interaction between a plant and a pathogenic bacterium, more than a hundred of the plant and of bacterial genes are now known to be involved. Most of these genes are either up-regulated or down-regulated during the disease development in compatible plants or during elicitation of resistance response in incompatible plants. Thus, the true understanding of the pathogenicity requires the information not only on the list of the structural genes involved in pathogenesis including suppression of disease resistance response but also on the regulatory mechanisms for their expressions. From such dynamic studies of bacterial genes, the highly sophisticated regulatory net works using both the pathogenicity-specific regulatory genes and various global regulatory genes are required for the unique expression of the pathogenicity-related genes. Here, we will focus on the findings from the dynamic analyses of the genes required for the pathogenic bacteria especially to enter into the disease life cycle in both wide and narrow host-range pathogens. We believe the understanding of these mechanisms should lead us to explore the new strategies to protect the plants at least from the infections by these pathogenic bacteria and most likely from those by other pathogens too.

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I-2

Participation of Host-Specific Toxins in Pathogenicity of Corynespora cassiicola. Fumihiro Nakano , Tetsuya Maruyama , Rox1

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ana Yanira Parada , Yoshifumi Shimomoto , Akinori Kiba , Yasufumi Hikichi , Noriyuki Miyake , Motoichiro Kodama and Hiroshi Otani . 1

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Faculty of Agriculture, Tottori University, Tottori 680-8553, Japan; Kochi Prefectural Agricultural Research Center, Nankoku 783-0023, Japan; Faculty of Agriculture, Kochi University, Nankoku 783-8502, Japan; Aichi Agricultural Research Center, Nagakute 480-1103, Japan 1

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Corynespora cassiicola is known as the pathogen, which has a wide host range. Recently, C. cassiicola, which has specialized host range has been also reported. We reported that C. cassiicola causing Corynespora target spot of tomato and C. cassiicola causing spot of

cucumber are pathogenic to respective host, and produce host-specific toxins (HSTs) named CCT-toxin and CCC-toxin, respectively. In 2004, a new disease occurred on leaves, stems and fruits of sweet pepper in Kochi prefecture in Japan, and the pathogen was identified as C. cassicola causing Corynespora blight of sweet pepper (sweet pepper pathogen). Spore inoculation test showed that the pathogen is pathogenic to eggplant and tomato in addition to sweet pepper. In 2007, moreover, a new disease occurred on leaves of Mandevilla, an ornamental plant, in Aichi prefecture in Japan, and the pathogen was identified as C. cassiicola causing Corynespora leaf spot of Mandevilla (Mandevilla pathogen). In spore inoculation test, pathogenicity of the pathogen was specific to Mandevilla. Therefore, culture filtrates (CFs) of sweet pepper and Mandevilla pathogens were assayed for toxicity. The CF of sweet pepper pathogen induced necrosis on leaves of sweet pepper, eggplant and tomato, but not on leaves of non-host plants. When the CF was subjected to DIAION HP-20 column chromatography, the unbound fraction caused necrosis on leaves of sweet pepper, eggplant and tomato, and the fraction eluted with 50% methanol had toxicity on tomato leaves in same fraction as that of CCTtoxin. These results indicate that sweet pepper pathogen may produce at least two HSTs, one is active to sweet pepper, eggplant and tomato, and the other is same or similar to CCT-toxin. The CF of Mandevilla pathogen had specific activity on Mandevilla, indicating that the pathogen may produce HST active to host plant. Purification of these toxins is now in progress.

I-3

Production of a Host-Specific Protein Toxin by Alternaria brassi-

cae. Roxana Y. Parada , Kumiko Oka , Daisuke Yamagishi , Emi Sakuno , Nobuhiro Mori , Mayumi Egusa , Motoichiro Kodama and Hiroshi Otani . Faculty of Agriculture, Tottori University, Tottori 6801

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8553, Japan; Agricultural, Food and Environmental Sciences Research Center of Osaka Prefecture, 583-0862, Japan 2

Leaf spot caused by Alternaria brassicae is a major disease in cultivated Brassica plants worldwide. Destruxin B, a cyclic peptide produced by A. brassicae, has been reported to be a host-specific toxin (HST) as agents of virulence or pathogenicity. However, we found that host specific toxicity is detected in high molecular weight fraction (HMW, >5 kDa) from spore germination fluid (SGF), but not in low molecular fraction (LMW,