Cucumber mosaic virus (CMV) belongs to genus Cucumovirus. The Cucumovirus group contains three distinct members： CMV, Tomato aspermy virus (TAV), and Peanut stunt virus (PSV). The type member, CMV is the most widespread and most studied. CMV is isometric particles about 30 nm in diameter. The genome of CMV is divided into three RNAs. In addition, RNA extracted from virus particles contains a fourth RNA that is a subgenomic RNA generated from RNA3. RNA1 and RNA2 are each encapsidated in separate particles, whereas RNAs3 and 4 are coencapsidated in a third particle. Hence, inoculation by three particles, transmitted either mechanically or by the aphid vector, is required to infect plants.(중략)

Clubroot disease of curcifer crops caused by Plasmodiophora brassicae had been first reported in 1920 in Korea, and maintained mild occurrence until 1980s. Since 1990s the disease has become severe in alpine areas of Kyonggi and Kangwon, gradually spread to plain fields throughout the country, and remains as the greatest limiting factor for its production. Researches on the disease has begun in late 1990s in our laboratory after experiencing severe epidemics. Survey of occurrence and etiological and ecological studies have been carried out, particularly, on the pathogen physiology, race identification, quantification of soil pathogen population, host spectrum of the pathogen, and control measures.(중략)

When plants are infected by plant pathogens, rapid cell responses are initiated for further inhibition from fast invasion of pathogens. Hypersensitive response (HR) of plant is well known defense response stopping pathogenesis process through rapid cell death. However, informations on the signaling pathway from reception of pathogen by host plants to appropriate resistant responses are very limited to date. Efficient perception of infection by pathogens and well-programmed signalling mechanism for appropriate responses are important for survival of plants. Plant have developed a sophisticated network(s) of defense/stress responses, among which one of the earliest signalling pathways after perception (of stimuli) is the evolutionary conserved Rop GTPase and mitogen-activated protein kinase (MAPK) cascade.(중략)

Potato virus X (PVX), the type member of Potexvirus genus, is a flexuous rod-shaped virus containing a single-stranded (＋) RNA. Infection by PVX produces genomic plus- and minus-strand RNAs and two major subgenomic RNAs (sgRNAs). To understand the mechanism for PVX replication, we are studying the cis- and/or trans-acting elements required for RNA replication. Previous studies have shown that the conserved sequences located upstream of two major sgRNAs, as well as elements in the 5' non-translated region (NTR) affect accumulation of genomic and sg RNAs. Complementarity between sequences at the 5' NTR and those located upstream of two major sgRNAs and the binding of host protein(s) to the 5' NTR have shown to be important for PVX RNA replication. The 5 NTR of PVX contains single-stranded AC-rich sequence and stem-loop structure. The potential role(s) of these cis-elements on virus replication, assembly, and their interaction with viral and host protein(s) during virus infection will be discussed based on the data obtained by in vitro binding, in vitro assembly, gel shift mobility assay, host gene expression profiling using various mutants at these regions.

Contemporary biological control system includes the use of fungi to control weeds in agricultural ecosystems and forests. Fungal pathogens of weeds that are highly virulent and specific to target weeds, and able to be produced massively by artificial culture could be applied like chemical herbicides over the weeds.(중략)

Helicobasidium mompa Tanaka and Resellinia necatrix Prillieux cause violet root rot and white root rot of various crops, respectively. Intensive cultural practices, such as the use of dwarf stock, glasshouse cultivation, etc., predispose plants to the diseases. The diseases can be controlled only by biennial drench of 50100L of chemicals for each tree. Biocontrol with soil microorganisms proved ineffective under field conditions. Long-term control may be hampered by the perennial growth of hosts and by the difficulty in the establishment of antagonists in soil. Crop rotation or soil amendment is not applicable, either. Fungal viruses with dsRNA genome (Buck 1986) are promising against root diseases of fruit trees since they exist within the cytoplasm of fungal hyphae and need no effort to help them persist in the field. The viruses are considered to spread though the network of fungal mycelia in the soil once they enter the fungal cytoplasm. Here, we present preliminary results from a project to control the root diseases of fruit trees with dsRNA.(중략)

Fungicide treatment is the most important method for the control of plant diseases caused by phytopathogenic fungi. But fungicide resistant strains have appeared in many phytopathogenic fungi. Until now, molecular mechanisms of fungicide resistance such as mutation of target protein, overproduction of target enzyme and detoxification of fungicide have been designated. Recently, it was demonstrated that active efflux of fungicides mediated by ATP-binding cassette (ABC) transporters also contributes to fungicide resistance in several filamentous fungi, such as Aspergillus nidulans, Penicillium digitatum and Botrytis cinerea.(중략)

Chestnut blight, caused by Cryphonectria parasitica, is the most destructive disease of American and European chestnut trees. A total of 672 C prasitica was isolated from blight lesion on chestnut twigs, which were collected from major chestnut plantations all over Korea in 1999. Isolation rates were over 30% in Kyunggj-, Kyongnam-, and Chonnam-do. The highest isolation rate was 37.4% and recorded in Kyongnam-do. On the other hand, Chonbuk-do had the lowest isolation rate as 13.5%. In grouping of C parasitica by colony shape and color, yellow colony with irregular margin were the most dominant colony type with a frequency of 65.2%. When the 672 isolates were inoculated on the chestnut twigs, 380 isolates (56.5%) caused lesions larger than the standard virulent isolate EP155-2, while 158 isolates (23.4%) caused smaller lesions than the standard hypovirulent isolate UEP-1. In Bavendamm test that determines phenol oxidase activity, 97.1% of all the isolates resulted the same or darker discoloration than EP155-2, and only 12.2% resulted the same or lighter discoloration than UEP-1. In the vegetative compatibility (VC) tests, total 670 isolates were divided into 121 VC groups (VCGs). Kyongnam-, Chonnam-, and Chungnam-do, the three principal chestnut plantation area, had 49, 33, and 27 VCGs, respectively. Among the VCGs, the biggest VCG, KR-VC104, was composed of 164 isolates and the second biggest VCG had 62 isolates. But, 64 of 121 VCGs consisted of sole member. More than 65.8% of KR-VC104, was isolated from the three provinces, Kyongnam-, Kangwon-, and Chungbuk-do. In KR-VC104, 62.8%, 59.1%, and 85.9% of the isolates looked like virulent in colony type, pathogenicity test, and Bavendamm test. In ds-RNA detection tests using cellulose chromatography, 77 of total 650 isolates were ds-RNA positive and detected ds-RNA segments were approximately 12kb, 3kb, 2.7kb, 2kb, and 1.8kb in size. Among the 77 isolates, 46 isolates had 12kb and 25 isolates had 12kb and 2.7kb. Other 6 Isolates had small ds-RNA segments. Kyongnam-, Chonnam-, and Chungnam-do had 43, 16, and 5 ds-RNA positive isolates, respectively. Among the 121 VCGs, only 29 VCGs had ds-RNA positive isolates.(중략)

Since Rice bacterial grain rot (RGBR) was reported at 1986 in Korea, it has been severely occurred in 1994, 1995, 1998, and especially around 16,609 ha in 2000, and became a major disease in rice cultivation field. This study was focused on investigation of ecology of RGBR, weather conditions that affect development of epidemics, and development of an effective RGBR forecast system based on weather conditions during the rice heading period.(중략)

Since 1993, a total of 50 problematic plant diseases unrecorded in Korea were surveyed in Gyeongnam province. Totally 34 new host plants to corresponding pathogens investigated in this study were 5 fruit trees, 9 vegetables, 12 ornamental plants, 3 industrial crops, and 5 medicinal plants. Among the newly recorded fruit tree diseases, fruit rot of pomegranate caused by Coniella granati and Rhizopus soft rot of peach caused by Rhizopus nigricans damaged severely showing 65.5% and 82.4% infection rate. Among the vegetable diseases, corynespora leaf spot of pepper caused by Corynespora cassiicola and the crown gall of pepper caused by Agrobacterium tumefaciens, powdery mildew of tomato caused by Oidiopsis taurica were the most severe revealing 47.6%, 84.7%, and 54.5% infection rate in heavily infected fields, respectively. In ornamental plants, collar rot of lily caused by Sclerotium rolfsii, gray mold of primula caused by Botrytis cinerea, soot leaf blight of dendrobium caused by Pseudocercospora dendrobium, sclerotinia rot of obedient plant caused by Sclerotinia sclerotiorum showed 32.7 to 64.8% disease incidence. On three industrial plants such as sword bean, broad bean, and cowpea, eight diseases were firstly found in this study. Among the diseases occurring on broad bean, rust caused by Uromyces viciae-fabae and red spot caused by Botrytis fabae were the major limiting factor for the cultivation of the plant showing over 64% infection rate in fields. In medicinal plants, anthracnose of safflower caused by Collectotrichum acutatum was considered the most severe disease on the plant and followed by collar rot caused by Sclerotium rolfsii.(중략)

The International Committee for Taxonomy of Viruses (ICTV), which was formed over 30 years ago, aims to develop a single, universal taxonomic scheme for all viruses or, in other words, "the classification of viruses and the assignment of names to taxa". Plant Virus taxonomy is in charge of Plant Virus Subcommittee, a substructure of the ICTV. The ICTV has been most successfully pursuing that aim and its mammoth 'Seventh Report' records details of the names it has collated and approved, and of the classification, it has devised. The current 7th ICTV report published in 2000 contains plant viruses of 951 species in 79 genera in 17 families, though 24 of the 79 genera are floating genera, that is, they are not included in any established families. Proposed name of new or existing viruses are vote for the accepted taxonomic proposals by ICTV Executive Committee meeting. The approved results have been published as the ICTV reports providing standard names and taxa of viruses all over the world. A number of new plant viruses have been identified or reclassified in the genus or species level, and new genera and families have been proposed.(중략)

Incompatible plant-pathogen interactions result in the rapid cell death response known as hypersensitive response (HR) and activation of host defense-related genes. To understand the molecular and cellular mechanism controlling defense response better, several approaches including isolation and characterization of novel genes, promoter analysis of those genes, protein-protein interaction analysis and reverse genetic approach etc. By using the yeast two-hybrid system a clone named Tsipl, Tsil -interacting protein 1, was isolated whose translation product apparently interacted with Tsil, an EREBP/AP2 type DNA binding protein. RNA gel blot analysis showed that the expression of Tsipl was increased by treatment with NaCl, ethylene, salicylic acid, or gibberellic acid. Transient expression analysis using a Tsipl::smGFP fusion gene in Arabidopsis protoplasts indicated that the Tsipl protein was targeted to the outer surface of chloroplasts. The targeted Tsipl::smGFP proteins were diffused to the cytoplasm of protoplasts in the presence of salicylic acid (SA) The PEG-mediated co-transfection analysis showed that Tsipl could interact with Tsil in the nucleus. These results suggest that Tsipl-Tsil interaction might serve to regulate defense-related gene expression. Basically the useful promoters are valuable tools for effective control of gene expression related to various developmental and environmental condition.(중략)

Total 291 isolates of Ampelomyces quisqualis were obtained from 32 species of powdery mildew fungi and a selected isolate, Ampelomyces quisqualis 94013 (AQ94013) effectively hyperparasitized 6 species of Sphaerotheca and one species of Erysiphe which cause serious damage on many important crops in Korea. Moreover, AQ94013 showed antagonistic effects against 12 major fungal plant pathogens as well. Results indicated that the present isolate is not a host specific hyperparasite and has a broad spectrum of biocontrol potential. Providentially, AQ94013 revealed resistance to a number of agrochemicals so as to be applied with the chemicals reciprocally.(중략)

Plants have evolved differently from animals having mobile activities. Thus, plants should have developed unique defense mechanisms against biotic/abiotic stresses to which plants are differently exposed, according to seasons. Most organisms have an conserved signaling network using mitogen-activated protein kinase (MAPK) cascade(s). The phenomenon implied that they are functionally very important in all organisms. In fact, they constitute one of the major components of signaling pathways involved in regulating a wide range of cellular activities from growth and development to cell death. Recently, complete MAPK cascade was first characterized in Arabidopsis from the receptor kinase (FLS2) through fellowing MEKKI -MKK4/MKK5-MPK3/MPK6-WRKY22/MRKY29 pathway. Whereas, MAPK cascade signaling pathway in monocot plant including rice (0ryza sativa L.), the most important of all food crops and an established monocot plant research model, MAPKinase kinase kinases (MAPKKK) of rice are the first upstream component of the MAPK cascade, but MAPKKK has been first identified and characterized in our lab and designated as, OsEDRl based on its homology with the Arabidopsis EDRI. The Arabidopsis EDRl was regarded as a negative regulator of defense response and the role of rice OsEDRl was analyzed. Transcriptional regulation of OsEDRl was detected under various stresses and immunoblotting analysis is going on to detect the level of OsEDRl protein in the mutants showing unique phenotype. We also introduced the constitutively active and the dominant negative forms of the OsEDRl for characterizing biological function.

Phytophthora capsici is a pathogen on several economically important crops including pepper. In pepper growing areas in Korea, Phytophthora blight caused by p. capsici has been considered as the most serious problem in pepper production. The Oomycete attacks the roots, stems, leaves and fruits of the plant. To understand the molecular mechanisms involved in the disease development, the genes expressed doting pepper p. capsici interaction were explored by analyzing expressed sequence tags (ESTs). A complementary DNA (cDNA) library was constructed from total RNA extracted from pepper leaves challenged with p. capsici for 3 days resulting in early stage of symptom development. The comprehensive analysis on the single pass sequencing of over 4000 randomly selected cDNA clones with contig assembly, unique gene extraction, sequence comparison, and functional categorizing will be presented with an emphasis on the genes involved in plant defense and pathogenicity during disease development of the pepper Phytophthora blight.

The effect of biological control agent Bacillus sp. (BAC03-3-1, BAC03-3-2, BAC02-4) on pre- and postemergence Sclerotinia rot of perilla (Perilla frutescens var. japonica) caused by Sclerotinia sclerotiorum was determined from greenhouse field trials. The ability of this antagonist to reduce germination of sclerotia of S. sclerotiorum was also evaluated. In the greenhouse, suspension of BAC03-3-1 application as root drench of perilla, which provided as little as 10$\^$7/ cells/ $m\ell$ per gram of soil, significantly increased plant stand in pathogen-infested soil over that in the untreated control. All three isolates reduced the germination of sclerotia of S. sclerotiorum in loamy sand soils in the greenhouse. In loamy sand amended with rice bran the sclerotial germination was inversely correlated (r = -0.79) with perilla stand in the greenhouse. However, a higher rate of bacterial suspension with rice bran(Ig dwt./100g soil) than that applied with bacterial suspensions only was necessary to achieve a comparable reduction in sclerotial germination. In field study, all three isolates added to soil to provide 10$\^$7/ cells/$m\ell$ per gram significantly prevented Sclerotinia rot (73-85％) after 35 days of growth. The isolate BAC02-4, BAC03-3-1 and BAC03-3-2 gave final stands of 65 to 75, 60 to 70, and 55 to 60％, respectively. The addition of rice bran(1 ％) to loamy sand in the field resulted in a 10-fold increase in propagule numbers of the three isolates within 10 days of application.

Several fungal isolates were isolated from diseased monochoria(Monochoria vaginalis, weed of paddy field), which has an resistance to sulfonyl urea(S.U.) herbicide, and were evaluated in the laboratory and greenhouse as potential mycoherbicide. Eight fungi, Alternaria sp., Colletotrichum sp., Curvularia sp., Paenicillium sp and etc. were observed in the isolates. Pathogenicity testing were done on the monochorias in the greenhouse. Monochorias were inoculated with suspensions containing conidia of each isolate at the rates of 1.0 ${\times}$ 10$\^$5/ conidia/ml and 0.1％ Tween 80 with hand-gun sprayer. Curvularia sp. and an unidentified fungal isolate caused 90-95％ mortality on the monochorias 15∼20 days after inoculation. However the other isolates induced slight symptom of disease on the monochorias. In the early stage of disease development sun-burn appearance was shown at the infected site and the last infected leaves and stems were withered to death. Subsequently the pathogenicity on the rice was evaluated with above two effective isolates. From the test an unidentified isolate showed pathogenicity on the rice but Curvularia sp., named as DBB2003, didn't. Now the mass production and formulation using Curvularia sp. DBB2003 are in progress and the field test will be followed. Combination product with Curvularia sp. DBB2003 and chemical herbicide will be more effect to control the monochoria resisted on S.U. herbicide and need to be further tested.

The occurrence of Fusarium wilt in strawberry fields in Korea was assessed from 2001 to 2003. Fusarium wilt was found from June to August in nursery beds, from September to October after planting in production beds, and from January to March during harvest. The symptoms seen were root rots, discolored vascular tissue in the crown and deformation and yellowing of central leaflets. The disease occurred in up to 30％ of plants in 37 of 214 fields surveyed. Fusarium of sporum Schlecht. ex Fr. f. sp. fragariae was frequently isolated from cvs. Dochiodome, Maehyang, Redpearl, Samaberry and Akihime. Factors affecting the occurrence of Fusarium wilt were investigated; infested soils had high salt concentrations, low pH, OM, average P2O5 and exchangeable. Fusarium wilt was more frequent following conventional basal fertilization than after non-nitrogen basal fertilization and more frequent following the use of NH4-N than after NO3-N.

Apple stem grooving virus (ASGV) belongs to Capillovirus and infects pome fruits. Transmission mode of ASGV is known by grafting and mechanical inoculation into susceptible hosts, not by any other natural vectors. But we have observed the spread of ASGV in the field without mechanical inoculation or grafting. Transmission seems to be occurred from tree-to-tree and tree-to-susceptible herbaceous plants along but not across ditches in the field. In order to ascertain this possibility, various fungi were isolated and cultured from ASGV-infected plants and 69 isolates were characterized. By means of RNA dot-blot hybridization and PCR analysis, 3 isolates were sorted out for further studies. The isolates were identified to Tataromyces sp. and belonged to Phenicillium by morphological characteristics and molecular markers. As an experimental host, 10 kidney beans (Phaseolus vulgaris) were screened and Kyunggi-5 was selected for virus amplification and symptom development. Kyunggj-5 infected by fungi which seemed to carry ASGV showed the typical disease symptoms and viral coat protein genes were detected from all tested plants. To confirm the Koch's rule, fungi cultured from inoculation origins of kidney bean were grown on PDA media and re-inoculated to hosts. The fungi isolated from inoculation origins induced the typical disease symptoms on hosts. However virus free fungi did not induce any symptom on the experimental hosts. This bioassay showed that these typical symptoms were caused by virus, not fungi.

Citrus canker is very important disease in international trade of citrus. The disease was usually take place from late of June, and severe middle of July to middle of August, though disease occurrence was affected by environmental conditions. In pathogenicity test, three varieties, orange, lemon and kiyomi among 7 varieties, were succeptible, two varieties, satsuma mandarin and iwasachi, intermediate resistant. On the other hand, shiranuhi and yuzu were resistant relatively. The pathogen, Xanthomonas axonopodis pv. citri, grew well in PD broth adjusted to pH 7.0 at 26$^{\circ}C$. It's growth was best in medium containing group of monosaccharide as a carbon source and group of ammonium as a nitrogen source. Tow isolates were resistant to streptomycin among 11 isolates isolated from diseased leaves in field in Jeju-Do. The streptomycin sensitives isolate was controlled by in greenhouse test. On the other hand, the resistant and sensitive isolates were controlled by treatment with copper sulfate, the control value is 88.7％ and 90.6％, respectively.

Incidence of crown gall on lower stem of chrysanthemum, Chrysanthemum morifolium Ramat., was first observed at Hwasung, Gyeonggi, Korea in 2001, Tumors on the stem were 1.5-2 cm in size and semi-round with rough surface texture of dark brown color. Four strains of bacteria isolated from the tumor tissues were characterized. Their colonies were convex, glistening, circular with an entire edge, and white to tannish-cream in color on PDA plus CaCO$_3$. They were gram negative, oxidase positive, and growing on DIM agar. The bacterial isolates inducing gall formation in chrysanthemum were identified as Agrobacterium tumefaciens based on biochemical and physiological characteristics, fatty acid profile using Sherlock Microbial Identification System, and substrate utilization patterns using Biolog Identification System. Young chrysanthemum plants inoculated with the bacteria developed typical galls within two to three weeks. Seedlings of tomato and slices of carrot roots also produced typical galls two to three weeks after inoculation. This is the first report on crown gall of chrysanthemum in Korea.

Gummy stem blight pathogen is very difficult not only to monitor the inoculum levels prior to host infection, and also it is destructive and hard to control in field condition. We have applied RAPD technique to elucidate the genetic diversity of the genomic DNA of Didymella bryoniae and also to generate specific diagnostic DNA probe useful for identification and detection. The 40 primers produced clear bands consistently from the genomic DNA of twenty isolates of Didymella bryoniae, and two hundred seventy-three amplified fragments were produced with 40 primers. The combined data from 273 bands was analyzed by a cluster analysis using UPGMA method with an arithmetic average program of NTSYS-PC (Version 1.80) to generate a dendrogram. At the distance level of 0.7, two major RAPD groups were differentiated among 20 strains. RAPD group (RG) I included 8 isolates from watermelon except one isolate from melon. RAPD group (RG) IV included 12 isolates from squash, cucumber, watermelon and melon.. In amplification experiment with SCAR specific primer RG1F-RG1R resulted in a single band of 650bp fragment only for 8 isolates out of 20 isolates that should be designated as RAPD Group 1. However, same set of experiment done with RGIIF-RGIIR did not result in any amplified product.. Our attempts to detect intraspecific diversity of ITS region of rDNA by amplifying ITS region and 17s rDNA region for 20 isolates and restriction digestion of amplified fragment with 12 enzymes did not reveal polymorphic band. In order to develop RAPD markers for RGIV specific primer, a candidate PCR fragment( ≒1.4kb) was purified and Southern hybridized to the amplified fragment RGIV isolates. This promising candidate probe recognized only RGIV isolates

Ammi majus (white lace flower, Unbelliferae) is an ornamental plant used for cut-flower arrangements worldwide. A potyvirus was isolated from its leaves with mosaic and chlorotic symptoms in the cultivated field of Chiba, Japan. Compared with Japanese homuort mosaic virus (JHMV) previously isolated from Cryptotaenia japonica, it showed similar characteristics in host reactions and molecular properties. The nucleotide sequences of coat protein and 3'- nontranslated region were highly homologous and shared 87％ and 91％ identities with those of JHMV, respectively. This virus was thus supposed to be an isolate of JHMV and designated as JHMV-Am. Phylogenetic tree was constructed using CP nucleotide sequences of the two isolates and other potyviruses previously reported. JHMV-Am and JHMV fell into a cluster with Korean strain of Zantedeschia mosaic virus (ZaMV-KR). However, low identity in amino acid sequences was found in the termini of CP genes between the two isolates of JHMV and ZaMV-KR.

Inoculation of primary pepper leaves with an avirulent strain of Xanthomonas campestris pv. vesicatoria induced systemic acquired resistance (SAR) in secondary leaves. This SAR response was accompanied by the systemic expression of defense-related genes, a systemic microoxidative burst generating H2O2, and the systemic induction of ion-leakage and callose deposition in the non-inoculated, secondary leaves. Some defense-related genes encoding PR-1, chitinase, peroxidase, PR10, thionin, defensin and zinc-finger protein were distiilctly induced in the systemic leaves. The systemically striking accumulation of H$_2$O$_2$and strong increase in peroxidase activity in pepper was suggested to contribute to the triggering of cell death In the systemic micro-HRs, leading to the induction of SAR. Treatment of non-inoculated, secondary leaves with diphenylene iodinium (DPI), an inhibitor of the oxidative burst, substantially reduced the induction of some defense-related genes and subsequently SAR.

Twenty-seven monogenic rice lines harboring major resistant gene for blast were screened to analyze their resistance spectrum to Korean blast fungus population using 190 isolates collected from 1985 to 2002. Especially, the monogenic line containing Pi-9 gene was screened using 320 isolates. Based on the monogenic lines-blast isolate interactions, the 27 rice lines were classified into 9 groups. The chinese rice cultivar LTH showed susceptible to all the tested isolates. Those lines IRBLz-Fu, ERBL5-M and IRBL9-W harboring Pi-z, Pi-5, and Pi-9, respectively showed broader spectrum of resistance than those rice lines having Pi-19, Pi-7 etc. Interestingly, the Pi-9 gene(IRBL9-W) showed resistance to most isolates collected before 2000, but it showed susceptible reactions to 5％ and 20％ of blast fungus population in 2001 and 2002, respectively. Population of virulent isolates to Pi-ta, Pi-b, and Pi-7 also were increased in 2002 compared to those before 2000.

In addition to the five well-characterized genes of Tobacco mosaic virus (TMV), this virus contains a sixth open reading frame (ORF6) that encodes a 4.8 kDa protein. TMV ORF6 overlaps the ORFs encoding the 30 kDa movement protein and the adjacent 17.5 kDa capsid protein. Although the 4.8 kDa protein could not be detected in vivo, alteration of the AUG codons of this ORF resulted in a mutant virus that attenuated the virulence of the mutated TMV in Nicotiana benthamiana, but not N. tabacum (tobacco). These sequence changes did not affect either the replication or movement of the mutated TMV. Expression of TMV ORF6 from the virus expression vector Potato virus X (PVX) intensified the virulence of this virus in N. benthmiana, but not tobacco, while expression of TMV ORF6 from the virus expression vector Tobacco rattle virus enhanced the pathogenicity observed in both N. benthamima and tobacco. Thus, the TMV ORF6 is a host- and virus-specific. virulence factor. However, two separate assays indicated that the TMV 4.8 kDa protein was not a suppression of RNA silencing. A fusion protein formed between the TMV 4.8 kDa protein and the green fluorescent protein was expressed from the PVX vector and localized to plasmodesmata. Possible roles of the 4.8 kDa protein in pathogenicity will be discussed

To protect the plant against several soil-borne pathogens, we are currently constructing disease-resistant transgenic root stock for the growth of cucurbitaceae vegetable plants, watermelon and gourd. We made a watermelon cDNA library from Cladosporium cucumerinum-Infected leaves for substractive hybriazation and differential screening. We isolated the several pathogen inducible cDNA clones, such as caffeoyl-CoA-methyltransferase, LAA induced protein, receptor-like kinase homolog, hydroxyproline-rich glycoprotein, catalase, calmodulin binding protein, mitochondrial ATPase beta subunit, methyl tRNA synthetase and WRKY transcription factors. We previously obtained CaMADS in pepper and galactinol synthase ( CsGolS) in cucumber that were confirmed to be related with disease-resistance. CaMADS and CsGolS2 were transformed into the inbred line 'GO701-2' gourd, the inbred line '6-2-2' watermelon and the Kong-dye watermelon by Agrobacterium tumerfaciens LBA4404. Plant growth regulators (zeatin, BAP and IAA) were used for shoot regeneration and root induction for optimal condition. Putative transgenic plants were selected in medium containing 100mg/L kanamycin and integration of the CaMADS and CsGO/S2 into the genomic DNA were demonstrated by the PCR analysis. We isolated major soil-borne pathogens, such as Monosporascus cannonballus, Didymella bryoniae, Cladosporium cuvumerinum from the cultivation area of watermelon or root stock, and successfully established artificial inoculation method for each pathogen. This work was supported by a grant from BioGreen 21 program, Rural Development Administration, Republic of Korea.

The aim of this study was to characterize the interactions of rice and Burkholderia glumae, a causal agent of bacterial grain rot of rice, at molecular levels using whole genomic sequences and to identify genes important for pathogenicity and symptom development. To do these, we sequenced whole genome of the bacterium and constructed cosmid clone profiles. We generated pools of mutants using various transposons and determined mutation sites by sequencing rescued plasmids. We focused on studying toxoflavin biosynthetic genes, quorum sensing regulation, and Hrp type III protein secretion systems. We found that two possible operons consisting of five genes are involved in toxoflavin biosynthesis and their expression is regulated by quorum sensing and LysR-type regulator, ToxR. We have isolated the nn PAI of B. glumae and characterized by mutational analyses. The hrp cluster resembled most the putative Type III secretion systems of B. pseudomallei, which is the causative agent of melioidosis, a serious disease of man and animals. The Hrp PAI core region showed high similarity to that of Ralstonia solanacearum and Xanthomonas campestris, however some aspects were dissimilar.

A gain-of-function mutant, SHM-11 obtained through gamma-ray mutagenesis, is resistant to rice blast caused by Magnaporthe grisea while wild type Sanghaehyanghyella is highly susceptible to the same disease. The resistance in the mutant was not race-specific when we tested with four races (KJ-201, KI-1113a, KI-313, KI-409) of M. grisea. To identify genes involved disease resistance in the gain-of-function mutant, genes specifically expressed in the mutant were selected by suppression subtractive hybridization using cDNAS of blast-inoculated mutant and wild type as a tester and a driver, respectively, Random 200 clones from the subtracted library were selected and analyzed by DNA sequencing. The sequenced genes represented three major groups related with disease resistance； genes encoding PR proteins, genes probably for phytoalexin biosynthesis, and genes involved in disease resistance signal transduction. A gene encoding a putative receptor-like protein kinase was identified as highly expressed only in the gain-of-function mutant after blast infection. The role of the putative receptor-like protein kinase gene during blast resistance will be further studied.

Phytophthora blight is a devastating disease of pepper and occurs almost anywhere peppers are grown. Phytophthora blight is caused by Phytophthora capsici and this pathogen can infect every part of the plant by moving inoculum in the soil, by infecting water on surface, by aerial dispersal to sporulating lesions. Management of Phytophthora blight currently relies on cultural practices, crop rotation, and use of selective fungicides. Since these treatments are a short-term management, a classical breeding for development of resistant pepper against the Phytophthora is an alternative. So far some of the resistant cultivars have been on the market, but those are limited regionally and commercially. Therefore, ultimately an elite line resistant against this disease should be developed, if possible, by biotechnology. We have set out a series of work recently in order to develop Phytophthora resistant pepper cultivar. For the first time, the cDNA microarray analysis was peformed using an EST chip that holds around 5000 pepper EST clones to identify genes responsive to Phytophthora infection. Total RNA samples were obtained from Capsicum annuum PI201234 after inoculating P. capsici to roots and soil and exposed to the chip. .Around 900 EST clones were up-regulated and down-regulated depending on the two RNA sample tissues, leaf and root. From those, we have found 55 transcription factors that may be involved in gene regulation of the disease defense mechanism. Further and in detail information will be provided in the poster.

A rice cultivar, Taebaegbyeo, is highly resistant to rice blast and moderately resistant to bacterial leaf blight (BLB) caused by Magnaporthe grisea and Xanthomonas oryzae pv. oryzae, respectively. To study the rice disease resistance mechanism, we generated rice deletion M3 mutants by gamma-ray irradiation. Blast and BLB responses of 16,000 M3 mutants were screened by inoculating mixtures of 4 races (KJ-201, H-1113a, KI-313, KI-409) of M. grisea and 3 Korean races of X. oryzae pv. oryzae. We selected so far 21 M3 mutants of Taebaegbyeo showing high susceptibility to the diseases. One of the mutants, KCT-6417, was susceptible to KI-1113a race of M. grisea, suggesting the deletion of a race-specific blast resistance gene in the mutant. To isolate rice genes involved in blast resistance and defense response, we take a PCR-based suppression subtractive hybridization approach using cDNAs of blast-inoculated wild type and the KCT-6417 as a tester and a driver, respectively. Genes specifically expressed in the wild type will be presented. The selected genes would give us a clue to understand mechanism for the race specific resistance and defense responses against M. grisea H-1113a in Taebaegbyeo.

When plants are infected by plant pathogens, typical disease symptom termed lesion, appears in compatible interaction. Whereas, in incompatible interactions, only small speck of lesions are visible on the leaf surfaces. Hypersensitive response (HR) of plant which is the result of infection by incompatible pathogens, is a well known defense response inducing rapid cell death resulting in complete resistance. However, some rice mutants show spontaneous disease symptoms during the growth stages without interaction with pathogens. We investigated the spontaneous cell death mutant called Blast Lesion Mimic(BLM) generated by EMS mutation, on the relationship with the hypersensitive response as well as resistant characteristics. Accumulation of phenolic compounds were detected around the lesions as lesions develop on leaf surface. Activation of PR gene was detected before the lesion appeared, and that result indicates the defense-related response are started earlier than lesion formation. The BLM mutant showed resistant response to inoculation of Magnaporthe grisea KJ201 with which the wild type Hwacheong is totally susceptible. Informations on the formation of spontaneous lesions and detail analysis of lesion mimic mutants and related genes are very limited to date. It is really important to understand the phenomenon of the defense-related lesion formation for developing resistant cultivar for rice blast pathogens

Genetic transformation carried out to induce the pathogenicity mutants from the two isolates, Elsinoe fawcettii R-34 and MUD of citrus scab fungus to hygromycin resistant by transferring plasmides (pUCATPH) that contain hygB gene. We produced protoplast for transformation by using of combinations of available enzymes including ${\beta}$-D-glucanase, ${\beta}$ -glucuronidase, Iyticase and driselase. The protoplasts regenerated at 64 $\mu\textrm{g}$/ml of hygromycin B but not 128 $\mu\textrm{g}$ in sensitivity test to identify the concentration of useful marker for the selection of transformants. Approximately 1200 and 67 hygromycin resistant isolates from strain R-34 and strain MUD, respectively, were isolated on PDA added with 200 $\mu\textrm{g}$ /ml of hygromycun B. Fifty seven and 4 of hygromycin resistant isolates from strain R-34 and MUD, respectively, did not produce necrotic lesions on the leaf in detached-leaf assay. Finally, 9 isolates were isolated from strain R-34, and these Isolates produced non or very few symptoms on seedlings of citrus in greenhouse pathogenicity test. And it's very interesting that some isolates produced melanose-like symptom on very young leaves which it was not typical symptom and somtimes produced on only expanded leaf.

RSH (relA/spoT homolog) has been known to determine the level of guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp), which are the effector nucleotide of the prokaryotic stringent response and also play a role in antibiotic production and differentiation in Streptomyces species but not a little in eukaryotic organism, especially in plant. Salicylic acid (SA), a critical signal molecule of establishing systemic acquired resistance (SAR), could induce SAR in Pepper (Capcicum annuum) against Phytophthora capsici. And the extent of SAR induction was in proportion to the dosage of SA (or BTH). Suppression subtractive hybridization (SSH), a PCR-based method for cDNA subtraction, was carried out between SA-treated and non-SA-treated pepper leaves to isolate genes which may be responsible for defense signaling against pathogens. Early upregulated gene was selected from reverse northern and kinetics of SSH-genes transcripts in SA-treated pepper leaves upon SA treatment. Full-length cDNA of the gene (PepRSH； Pepper RelA / SpoT homolog) had an open reading frame (ORF) of 2166 bp encoding a protein of 722 amino acids and a significant homology with (p)ppGpp phosphohydrolase or synthetase. Genomic DNA gel blot analysis showed that pepper genome has at least single copy of PepRSH. PepRSH transcripts was very low in untreated pepper leaves but strongly induced by SA and methyljasmonic acid (MeJA), indicating that PepRSH may share common SA and MeJA-mediated signal transduction pathway Functional analysis in E. coli showed PepRSH confers phenotypes associated with (p)ppGpp synthesis through a complementation using active site mutagenesis.

For the selection and breeding of chestnut varieties resistant to the chestnut blight fungus Cryphonectria parasitica, disease resistance and susceptibility of 28 varieties widely planted and growing in Korea were evaluated by artificial inoculation of a pathogenic fungus. For this experiment, a typical virulent strain (KCPC-19) was selected. Artificial inoculation was conducted into all varieties by using two different materials and methods, i.e., bark and wood tissue sections in the laboratory and living trees in the field. In the bark and wood tissue section method, the size of necrotic area and canker development on chestnut varieties were examined and compared 4 days after inoculation. There were wide variations of chestnut varieties in disease resistance and susceptibility against chestnut blight fungus, but 3 varieties, Daebo！, Ishizuchi, and Sandae, were shown to be relatively resistant to the disease with the necrotic area of 0.95-1.03 cm2, while Arima was the most susceptible with the size of 2.0 cm2. In the living tree inoculation examined 5 weeks after inoculation, 3 varieties, Daebo, Ishizuchi, and Riheiguri, showed the higher resistance, but Tono 2 did the highest susceptibility among tested varieties.

It was investigated whether culture filtrates produced by X. fastiduosa could be used to determine varietal susceptibility in grape cultivars to anthracnose as a substitute for pathogen inoculation or field screening. Bioassay of grape leaves with culture filtrates showed that their phytotoxicities were active and host-selective. Ethyl acetate extracts from those also showed the toxicities and host selectivity among grape cultivars. The sensitive range of plants to culture filtrates and their ethyl acetate extracts was consistent with the host range to the pathogen. Susceptible cultivars were sensitive to even highly diluted culture filtrates but resistant cultivars were not affected even at original culture filtrates. Susceptible cultivars were sensitive to the undiluted culture filtrates than highly diluted culture filtrates and the younger leaves were the more sensitive to the culture filtrates and their ethyl acetate extracts in grapes.

To evaluate the resistance to crown gall in grape rootstocks, cuttings from twenty seven grape rootstocks were inoculated with Agrobacterium vitis Cheonan 493 and size of galls from grapevines was measured in a greenhouse. Tumors were formed in all varieties of grape rootstocks tested in this study and no grape rootstock variety was immune to crown gall. Tumors were found on the stems of all plants tested in '196-17'and '41B' Based on measuring size and weight of galls formedon the stem of grape rootstocks, '779P' was extremely susceptible to crown gall. Some varieties such as 'Gloire', '140R', '101-l4M', '3309C', and '333EM' found to be resistant, while '99R', '1447P', 'Rupestris du lot', '110R', 'Freedom', and '41B'were susceptible and '1103P', '5C', '420A', 'Golia', and '5BB' were moderately susceptible to crown gall.

Most major plant pathogenic bacteria in Korea belong to Xanthomonas spp.. Xanthomonas oryzae pv. oryzae is a major pathogen in rice, X. campestris pv. vesicatoria in pepper, X. axonopodis pv. giycines in soybean, X. campestris pv. campestris in cabbage, and X. axonoposid pv. citri in tangerin. Host specificity of the bacterial pathogen depends on the avirulence gene in the pathogen and the corresponding resistance gene in host plants. Many avirulence genes in bacteiral pathogen located on the native plasmids. However, the presence of the native plasmids in Xanthomonas spp. was not investigated well. In order to study the host specificity, we isolated native plasmids from Xanthomonas spp. and compared those plasmids each other, The presence of the native plasmids and the characteristics of the plasmids depended on the bacterial strains. In the X. axonopodis pv. glycines, most strains carried native plasmids but some strains did not. Some strains carry about 60 kb native plasmids including 3 different aviurlence genes. We will discuss the characteristics of the native plasmids isolated from the Xanthomonas spp.

Ralstonia solancearum infects many solanaceous plants, however race 3 infects only potato and tomato weakly. To identify genes responsible for race specificity of R. solanacearum, we mobilized genomic library of LSD2029 (race 3) into LSD341 (race 1) and inoculated 1,000 transconjugants into hot pepper. One transconjugant that did not induce wilt symptom in hot pepper was isolated. We found that a cosmid clone, pRSl, conferred avirulence to LSD341. By deletion and mutational analyses of pRSl, we found the 0.9-kb PstI/Hindlll fragment carries avirulence functions. We sequenced the fragment and identified one possible open reading frame, a rsal gene, possibly encoding 110 amino acids. The rsal was preceded with a plant-inducible promoter (PIP) box, indicating that the gene might be regulated by HrpB. Interestingly, the promoter region of the rsal homolog in the strain GM11000 (race 1) did not have the PIP box. Rsal did not show any significant homologies with proteins in the database, indicating th e protein is different from the previously reported avirulence proteins. When we mutated the rsal gene by marker-exchange in LSD2029, the mutant was less virulent in potato.

Generally, plants defend themselves against pathogens by structural and biochemical reactions. Defense structures act as physical barriers and inhibit the pathogen from gaining entrance and spreading through the plant. Xanthomonas axonopodis pv glycines, the causal pathogen of bacterial pustule of soybean, causes hypersensitive response (HR). When pepper fruits were inoculated with X. axonopodis pv. glycines, in situ, time-series defense-related structural changes occurred in the inoculated sites. Early responses were programmed cell death (PCD), characterized by condensation and vacuolization of the cytoplasm, condensation of nuclear materials, and fragmentation of the nuclear DNA, which were observed by transmission electron microscopy. Nuclear fragmentation was proven by TUNEL method under confocal laser scanning microscopy and DNA laddering through eletrophoresis. At later stages, plant responses were cell elongation and cell division, forming a periderm-like boundary layer that demarcated healthy tissues from the inoculation sites. Using several stains such as toluidine blue, sudan IV, annexin V, and phloroglucinol-HCl, defense-related materials and structural changes were also examined.

We found a soybean (Glycine max) cultivar 561 that was strongly resistant to a virulent bacterial strain of a Pseudomonas spp. Further identification revealed that the Pseudomonas spp. was a strain of Pseudomonas aeruginosa. Furthermore we identified specific genes involved in the resistance of soybean 561 and analyzed the pattern of gene expression against the Pseudomonas infection using differential-display reverse transcription PCR (DDRT-PCR). More than 126 cDNA fragments representing mRNAs were induced within 48 hours of bacteria inoculation. Among them, 28 cDNA fragments were cloned and sequenced. Twelve differentially displayed clones with open reading frames had unknown functions. Sixteen selected cDNA clones were homologous to known genes in the other organisms. Some of the identified cDNAs were pathogenesis-related genes (PR genes) and PR-like genes. These cDNAs included a putative calmodulin-binding protein, an endo-1,3-1,4-b-D-glucanase, a b-1,3-endoglucanase, a b-1,3-exoglucanase, a phytochelatin synthetase-like gene, a thiol pretense, a cycloartenol synthase, and a putative receptor-like sorineithreonine protein kinase. Among them, we found that four genes were putative pathogenesis-related genes (PR) induced significantly by the p. aeruginosa infection. These included a calmodulin-binding protein gene, a b-1,3-endoglucanase gene, a receptor-like sorine/threonine protein kinase gene, and pS321 (unknown function). These results suggest that the differentially expressed genes may mediate the strong resistance of soybean 561 to Pseudomonas aeruoginosa.

In order to cultivate watermelon on farm, grafting of the watermelon seedling to the watermelon stock is necessary because the watermelon root is less viable than the root of watermelon stock. Recently, commercially important watermelon varieties further require a resistant stock against especially CGMMV to control the heavy loss of the total yield of watermelon by CGMMV infection. Therefore, we have set out a project to develop a CGNEMV-resistant watermelon stock. We have successfully transformed dozens of watermelon stocks (gongdae) during last two years especially using a cDNA encoding the coat protein of CGMMV (cucumber green mottle mosaic virus). Recently we have tested levels of resistance of those watermelon stocks against CGMMV infection. For CGMMV inoculation, the leaves of one month old gongdae (T1) were rubbed by carborundum mixed with the CGMMV. A total of 140 plants (T1) were exposed to the CGMMV and we found that ten plants were completely resistant to virus infection. This is the first report that by genetic engineering a cucubitaceae crop resistant to CGMMV infection is ever developed. Further information will be provided in the poster.

Typical whiches broom symptoms caused by phytoplasma were observed in Ash (Fraxinus rhynchophylla Hence) in Korea. The symptoms were showing abnormally small leaves, short internodes, and proliferation of shoots. Fluorescence and electron microscopy of leaf midribs revealed phytoplasma positive DAPI fluorescence and numerous phytoplasma bodies localized in the phloem sieve tubes. Phytoplasma DNA of 1.8 Kb was detected consistently from all symptomatic samples by the amplification of phytoplasma DNA with the phytoplasma specific primer pair Pl/P7. But no phytoplasma DNA was detected in healthy ash seedlings. Based on sequence analyses of an amplified region, this phytoplasma is closely related to Eqilodium phyllody, Mulberry dwarf, and Aster yellows phytoplasmas with the homology of 99.95 ％, 99.79 ％ and 99.78 ％, respectively, This phylogenetic analyses indicate that ash witches broom phytoplasma but is evidently distinct from the ash yellows group 16SrⅦ and should be classified into the Aster yellows group 16SrⅥ.

Differential display (DD) of mRNA is a technique in which mRNA species expressed by a cell population are reverse transcribed and then amplified by many separate polymerase chain reactions (PCR). Using DD-RT-PCR we obtained many genes that expressed differentially in healthy and PVX-infected Nicotiana benthamima, using total RNAs extracted from healthy and PVX-infected N. benthamiana plants. Three hundred and twenty-five DNA fragments isolated from DD-RT-PCR were cloned and sequenced for further characterization. Several host genes including SKPI-like protein, heat shock transcription factor and Avr9/Cf-9 rapidly elicited protein were selected to obtain full-length open reading frame and to characterize their potential involvement in virus disease development and/or host's defense against virus infection employing PVX-based expression vector. Transcrips from wild-type and clones containing each selected gene were inoculated onto N. benthamiana Levels of virus replication were confirmedby RT-PCR and RNA blot analysis, Expression profiles and potential role(s) of selected genes upon PVX infection will be discussed.

Arabidopsis infected with beet curly top virus (BCTV) has the systemic symptoms like stunting of Plant growth, curling of leaves and shoot tips, and callus induction. The regulation of sucrose metabolism by BCTV infection is essential for obtaining the energy source in the process of virus replication and symptom development. Sucrose metabolism-associated gene expression and biochemical enzyme activity were analyzed with the rossette leaves and inflorescencestems of BCTV infected Arabidopsis by the time course of 1, 7, 14, 21 day postinoculation. The expression of invertase and sucrose synthase genes ( encoding sucrose-cleaving enzymes )was increased and reversely the level of Atkin10a ( sucrose non-fermenting gene ) was decreased, resulting by semi-quantitative reverse transcription polymerase chain reaction. The biochemical analysis of invertase and sucrose synthase activity was performed. The activity of neutral invertase in the inflorescence stems was elevated remarkably. The photosynthetic response in the source of sucrose metabolism was consistent with the down-regulation of ribulose 1,5 bisphosphate carboxylase gene, and lower activity than mock-inoculated plants. The levels of genes pertaining to the cell cycle, hormone, and biotic stress-related pathway showed an increase or a decrease dependent on viral symptoms. Therefore, sucrose sensing by BCTV infection can regulate the expression of sucrose metabolism-related key enzymes such as invertase and Atkin10a, and these gene products might influence to symptom development.

Barely yellow mosaic(BaYMV) and barely mild mosaic (BaMMV) bymoviruses are both transmitted by the soil-inhabiting fungus Polymyxa gramnis, and are responsible for economic losses in barley crops in Asia and Europe. Because chemical control of the vector is ineffective, the losses can only be prevented by growing resistant barley cultivars. The objective of this study is to produce resistant barley plants by transformation with viral coat protein(cp) genes. Resistance tests of T1 plants transformed with the BaYMV CP gene showed that at least four independent lines had clear resistance to BaYMV but two other lines were highly susceptible with severe symptoms. The CP gene was detected in all resistant T1 plants by genomic PCR. Most of T2 progenies derived from the resistant T1 lines also showed resistance. In contrast, only one out of 21 independent T2 lines transformed with the BAMMV CP gene tested showed clear resistance to BaMMV, and others were very susceptible. Further analyses of resistance and CP gene expression are in progress.

To develop an effective protection strategy against tomato bushy stunt virus (TBSV), tobacco plants expressing single-chain Fv antibodies (scFv), were established. A previous had shown that the replication activity of viral replicase was inhibited by the selected scFvs. Moreover, no systemic symptom was found after virus inoculation on leaves of wt N. benthamiana infiltrated with an Agrobacterium suspension resulting i3l expression of the scFvs. However, control plants showed systemic symptoms. In this study the localization of the scFvs within two transgenic plant lines, (CP28H3, CP-P55) was demonstrated using immunogold labelling. The gold particles, indicating the presence of scFv, were mostly found In the cytoplasm of the plant cells including chloroplasts and in the cell walls. However, they were hardly found in the vacuole, nucleoplasm and intercellular spaces. Gold particles often accumulated in either the cytosol or chloroplasts showing a specific labeling, There was no difference in type of gold labeling between both transgenic lines. The localization of the scFv in the cytoplasm further conforms the inhibition of the RNA-dependent RNA polymerase (RdRp) by the selected scFv because it is known that the RdRp is localized to membraneous cytosolic structures.

The coat protein (CP) of Turnip crinkle virus (TCV) is organized into 3 distinct domains, R domain (RNA-binding) connected by an arm, 5 domain and P domain. We have previously shown that the CP of TCV strongly suppresses RNA silencing, and have mapped N-terminal R domain of which is also the elicitor of resistance response in the Arabidopsis ecotype Di-17 carrying the HRT resistance gene. In order to map the region in the TCV CP that is responsible for silencing suppression, a series of CP mutants were constructed, transformed into Agrobacterium, coinfiltrated either with HC-Pro (the helper component proteinase of tobacco etch potyvirus) known as a suppressor of PTGS or GFP constructs into leaves of Nicotiana benthmiana expressing GFP transgenically. In the presence of HC-Pro, all CP mutants were well protected, accumulating mutant CP mRNAs and their proteins even 5 days post-infiltration (DPI). In the presence of GFP, some mutant constructs which showed the accumulation of CP mutants and GFP mRNAs at early stage but eventually degraded at 5 DPI. Only a mutant which carrying 4 amino acid deletion of R domain was tolerable to maintain suppressing activity, suggesting that the suppressing activity is not directly related with the eliciting activity. A transient assay also revealed that the mutants synthesized their proteins, suggesting that a full length of CP sequences and its intact structure are required to stabilize CP, which suppresses the RNA silencing.

The complete nucleotide sequences of genomic RNA of an isolate of soybean mosaic virus (SMV-CN18), which has ability to overcome Rsv resistance of soybean, have been determined. A large open reading frame encodes a polyprotein of 3068 amino acids with a predicted Mr of 350 kDa. Based on comparison with the proposed cleavage site of other potyviral polyproteins, nine mature proteins are predicted as a following order, P1, HC-Pro, P3, CI, 6K, VPg, NIa, NIb and coat protein (CP). The mature proteins of the strain share various amino acid identity with known SMV-G2, -G7 and -N strain, with the greatest variability occurring in the P1 (91 ％, 88 ％, 96％)and the lowest variability in the CP (100 ％, 99 ％, 100 ％). In addition, 5' untranslated region determined by 5' RACE is much more various than any coding regions. Difference in amino acid sequences throughout the genome is discussed in relation to resistance and susceptibility of soybean cultivars to SMV-CNl8.

We have isolated a full-length cDNA clone, CaCDPK4 encoding a typical calcium-dependent protein kinase (CDPK) from hot pepper cDNA library. Genomic southern blot analysis showed that it belongs to a multigene family, but represents a single copy gone in hot pepper genome. RNA expression pattern of this gene revealed that it is induced by infiltration of Xanthomonas axonopodis pv. glycines Bra into hot pepper leaves but not by water deficit stress. However, high salt treatment of NaCl (0.4 M) solution to hot pepper plants strongly induced CaCDPK4 gene. In addition, this gene is weakly responsive to the exogenous application of salicylic acid or ethephon. Biochemical study of the GST-CaCDPK4 recominant protein showed that it autophosphorylates in vitro and the presence of EGTA, a calcium chelater, eliminates the kinase activity of the recombinant protein. As a way to identify the in vivo function of CaCDPK4 in plants, VIGS (Virus-Induced Gene Silencing) was employed. Agrobacterium-mediated TRV silencing construct containing the kinase and calmodulin domain of CaCDPK4 resulted in cell death of Nicotiana benthamiana plants. A highly homologous H benthamiana CDPK gene, NbCDPK5, to CaCDPK4 was cloned from N. benthamiana cDNA library. VIGS of NbCDPK5 also resulted in cell death. The molecular characterization of this cell death phenotype is being under investigation.

An EREBP/AP2-type transcription factor (CaPFl) was isolated by DDRT-PCR following inoculation of soybean pustule pathogen Xanthomonas axonopodis pv. glycines Bra which induces HR on pepper leaves. Genomic Southern blot analysis revealed that the CaPFl gene is present as a single copy within the hot pepper genome. The deduced amino acid sequence of CaPFl has two potential nuclear localization signals, a possible acidic activation domain, and an EREBP/AP2 motif that could bind to a conserved cis- element present in promoter region of many stress-induced genes. The mRNA level of CaPFl was induced by both biotic and abiotic stresses. We observed higher-level transcripts in resistance-induced pepper tissues than diseased tissues. Expression of CaPFl is also induced upon various abiotic stresses including ethephon, MeJA, cold stress, drought stress and salt stress treatments. To study the role of CPFI in plant, transgenic Arabidopsis and tobacco plants which express higher level of pepper CaPFl were generated. Global gene expression analysis of transgenic Arabidopsis by cDNA microarray indicated that expression of CaPFl in transgenic plants affect the expression of quite a few GCC box and DRE/CRT box-containing genes. Furthermore, the transgenic Arabidopsis and tobacco plant, expressing CaPFl showed tolerance against freezing temperature and enhanced resistance to Pseudomonas syrnigae pv. tabaci. Taken together, these results indicated that CaPFl is a novel EREBP/AP2 transcription factor in hot pepper plant and it may has a significant role(s) in regulation of biotic and abiotic stresses in plant.

An osmotin-like protein (CAOSMl) gene was isolated from pepper leaves infected with the avirulent strain Bv5-4a of Xmthomonas campestris pv. vesicatoria. The cDNA encodes a polypeptide of 250 amino acids with a molecular mass of 27, 361 Da. Its amino acid sequence is highly homologous to various osmotin-like proteins from other plant species. The CAOSMl gene expression was organ- and tissue-specifically regulated In pepper plants. The CAOSMl mRNA was intensely localized in the endodermis area of root tissue and in the phloem cells of vascular bundles of red fruit tissue, but not in leaf, stem, and green fruit tissues of healthy pepper plants. Infection by X. c. pv vesintoria, Colletotrichum coccodes, or Phytopkhora capsici iinduced CAOSMl transcription in the leaf or stem tissues. Expression of the CAOSMl gene was somewhat higher in the incompatible than the compatible interactions of pathogens with pepper. The CAOSMl mRNA was prevalently localized in the phloem cells of the vascular bundle of leaf tissues infected by C. coccodes. The CAOSMl gene was activated in leaf tissues by treatment with ethylene, methyl jasmonate, high salinity, cold acclimation and mechanical wounding, but not by abscisic acid (ABA) and drought. These results indicate that the pepper CAOSMl protein functions in response to Pathogens and some abiotic stresses in pepper plants

Pepper defensin ( CADEFl) clone was isolated from cDNA library constructed from pepper leaves infected with avirulent strain Bv5-4a of Xanthomonu campestris pv. vesicatoria. The deduced amino acid sequence of CADEFl is 82-64％ identical to that of other plant defensins. Putative protein encoded by CADEFl gene consists of 78 amino acids and 8 conserved cysteine residues to form four structure-stabilizing disulfide bridges. Transcription of the CADEF1 gene was earlier and stronger induced by X campestris pv. vesicatoria infection in the incompatible than in the compatible interaction. CADEF1 mRNA was constitutively expressed in stem, root and green fruit of pepper. Transcripts of CADEFl gene drastically accumulated in pepper leaf tissues treated With Salicylic acid (SA), methyl jasmonate (MeJA), abscisic acid (ABA), hydrogen Peroxide (H$_2$O$_2$), benzothiadiazole (BTH) and DL-${\beta}$-amino-n-butyric acid (BABA). In situ hybridization results revealed that CADEF1 mRNA was localized in the phloem areas of vascular bundles in leaf tissues treated with exogenous SA, MeJA and ABA. Strong accumulation of CADEF1 mRNA occurred in pepper leaves in response to wounding, high salinity and drought stress. These results suggest that bacterial pathogen infection, abiotic elicitors and some environmental stresses may play a significant role in signal transduction pathway for CADEF1 gene expression.

Two pathogen-induced hot pepper transcription factors (CaNACl and CapIfl) were introduced into‘MicroTom’tomato by Agrobacterium tumefaciens-mediated transformation. We used to nptII containing kanamycin resistance gene as a selection marker. Both transformed and non-transformed plants were transferred to pot after rooting test in vitro. To approximate the levels of caNACl transcript in leaves of wild-type and transgenic plants, RNA blots were hybridized with double-stranded full-length CaNACl probe at moderate stringency, Although the relative signal strength for hybridization fluctuated among the samples on different blots, transgenic plant lines N-1, N-2 and N-3 consistently displayed increased levels of CaNACl transcript relative to other transgenic lines and wild-type plants. Of all the transgenic lines examined, line N-7 had the least amount of CaNACl transcript. Role of these transcription factors in pathogen defense will be examined by overexpression in tomato.

To better understand plant defense responses against pathogen attack, we identified the transcription factor-encoding genes in the hot pepper Capsicum annuum that show altered expression patterns during the hypersensitive response raised by challenge with bacterial pathogens. One of these genes, Ca1244, was characterized further. This gene encodes a plant-specific Type IIIA - zinc finger protein that contains two Cys$_2$His$_2$zinc fingers. Ca1244 expression is rapidly and specifically induced when pepper plants are challenged with bacterial pathogens to which they are resistant. In contrast, challenge with a pathogen to which the plants are susceptible only generates weak Ca1244 expression. Ca1244 expression is also strongly induced in pepper leaves by the exogenous application of ethephon, an ethylene releasing compound. Whereas, salicylic acid and methyl jasmonate had moderate effects. Pepper protoplasts expressing a Ca1244-smGFP fusion protein showed Ca1244 localizes in the nucleus. Transgenic tobacco plants overexpressing Ca1244 driven by the CaMV 355 promoter show increased resistance to challenge with a tobacco-specific bacterial pathogen. These plants also showed constitutive upregulation of the expression of multiple defense-related genes. These observations provide the first evidence that an Type IIIA - zinc finger protein, Ca1244, plays a crucial role in the activation of the pathogen defense response in plants.

Several plant and microbial genes that could confer disease resistance in transgenic rice plants are being cloned and characterized. We are currently constructing transgenic rice lines that overexpress the gene products, such as a galactinol synthase, a defensin, and a bacterial ACC deaminase. Subtractive hybridization of a rice cDNA library constructed from the Xanthomonas oryzae-infected ice leaves resulted in isolation of many inducible cDNA clones including a elongation factor EF2, a oryzain alpha, a catalase, a aldehyde dehydrogenase, a S-adenosylmethionine synthetase, a caffeic acid O-methyltransferase, a glyceraldehyde-3-phosphate dehydrogenase, a light-regulated protein, nKY transcription factors, and a nucleotide diphosphate kinase. Some genes among those may be useful genetic sources for construction of disease resistant transgenic rice. Full lengths of the rice OsFIERG and a rice oryzain genomic clones were cloned, and serial deletion fragments of the promoter regions of these genes were fused with GUS reporter gene in pCAMBIA1201, respectively. Promoter activities of these constructs will be examined upon various stresses and Pathogen infections to obtain the pathogen specific inducible-promoter. This work was supported by a grant from BioGreen 21 Program, Rural Development Administration, Republic of Korea.

Since hot peppers (Capsicum annuum L.) are getting reputation as an important source of vitamins, medicine and many other areas, consumption and cultivation is being increased in the world. In spite of this usefulness, so little attention has been given to the hot pepper plants. To date, less than 500 nucleotide sequences including redundancy has been identified in NCBI database. Therefore we started to EST sequencing project for initial characterization of the genome, because of the large genome size of hot pepper (2.7 3.3 ${\times}$ 109 bp), To date, a set of 10,000 non-redundant genes were identified by EST sequencing for microarray-based gene expression studies. At present, cDNA microarrays containing 4,685 unigene clones are used for hybridization labeled targets derived from pathogen infected and uninoculated leaf tissues. Monitoring of gene expression profiles of hot pepper interactions with soybean pustule pathogen (Xag；Xanthomonas axonopodis pv. glycine) will be presented.

Although plants have evolved to possess various defense mechanisms from local biotic and abiotic stressors, most of yield loss is caused by theses stressors. Recent studies have revealed that several different stress responsive reactions are inter-networking. Therefore, the identification and dissection of stress responsive genes is an essential and first step towards understanding of the global defense mechanism in response to various stressors. For this purpose, we applied cDNA microarray analysis, because it has powerful ability to monitor the global gene expression in a specific situation. To date, more than 10,000 non-redundant genes were identified from seven different cDNA libraries and deposited in our EST database (http://plant.pdrs.re.kr/ks200201/pepper.html). For this study, we have built 5K cDNA microarray containing 4,685 unigene clones from three different cDNA libraries. Monitoring of gene expression profiles of hot pepper interactions with biotic stress, abiotic stresses and chemical treatments will be presented. Although this work shows expression profiling at the sub-genomic level, this could be a good starting point to understand the complexity of global defense mechanism in hot pepper.

Plants have evolved along with pathogens, and they have developed sophisticated defense systems against specific microorganisms to survive. G-protons are considered one of the upstream signaling components working as a key for the defense signal transduction pathway. For activation and inactivation of G-protein, GTP-biding proteins are involved. GTP -binding proteins are found in all organisms. Small GTP-binding proteins, having masses of 21 to 30kD, belong to a superfamily, often named the Ras supefamily because the founding members are encoded by human Ras genes initially discovered as cellular homologs of the viral ras oncogene. Members of this supefamily share several common structural features, including several guanine nucleotide binding domains and an effector binding domain. However, exhibiting a remarkable diversity in both structure and function. They are important molecular switches that cycle between the GDP-bound inactive form into the GTP-bound active form through GDP/GTP replacement. In addition, most GTP-binding proteins cycle between membrane-bound and cytosolic forms. such as the RAC family are cytosolic signal transduction proteins that often are involved in processing of extracellular stimuli. Plant RAC proteins are implicated in regulation of plant cell architecture secondary wall formation, meristem signaling, and defense against pathogens. But their molecular mechanisms and functions are not well known. We isolated a RacB homolog from rice to study its role of defense against pathogens. We introduced the constitutively active and the dominant negative forms of the GTP-hinging protein OsRacB into the wild type rice. The dominant negative foms are using two forms (full-sequence and specific RNA interference with RacB). Employing southern, and protein analysis, we examine to different things between the wild type and the transformed plant. And analyzing biolistic bombardment of onion epidermal cell with GFP-RacB fusion protein revealed association with the nucle.

Rice seedling disease is one of major problems in water-seeded rice. This disease is known to be caused by several pathogen such as Pythium, Achlya, and Fusarium species. However, seedling disease of rice in water-seeded rice in Korea is not extensively studied. Pythium species have been isolated from Seosan, Yeoju, Icheon areas using Pythium selective media and their pathogenicity was investigated. All of the Pythium isolates showed strong pathogencity causing seedling emergence reduction in water-seeded rice. Seedling emergence was reduced to 0∼9％ at 10 days after inoculation of 23 Pythium isolates compared to 60％ of noninoculated control in a growth chamber. However, Fusarium species did not cause seedling emergence reduction in water-seeded rice. In contrast, when no water added into water agar or soil, the pathogen caused seedling rot two weeks after planting. These results indicate that Pythium species is a cause of seedling disease in water-seeded cultivation areas in Korea.

A full length cDNA, OsLEUZIP, encoding leucine zipper containing protein from rice EST of rice (0ryza sativa L. cv. Dongjin) treated Xanthomonas oryzae pv. oryzae 10331. OsLEUZIP contains 1,227 bp nucleotides and encodes a protein of 408 amino acid residues with predicted molecular weight of 47,229 Da. The deduced amino acid sequence of OsLEUZIP has consensus sequence of leucine zipper from PROSITE (PDOC00029), L-X(6)-L-X(6)-L-X(6) -L. OsLEUZIP gene were preferentially induced in rice during incompatible interaction with Xanthomonas oryzae pv. oryzae 10331 and Pyracuraria grisea KJ-301. Expression of OsLEUZIP gene was also induced by treatment of abiotics such as ethephon and ABA. Our data represented in this study suggesting that OsLEUZIP gene may play an important role in the rice defense-related. Further studies of this gene, overexpression in rice, yeast-two hybrid assay, electrophoretic mobility shift assay and northern blot analyses of transgenic plant, would be useful to elucidate the role of the OsLEUZIP gene in defense responses of rice.

Magnaporthe grisea, the casual agent of rice blast, forms an appressorium to penetrate its host. Much has been learned about environmental cues and signal transduction pathways, especially those involving CAMP and MAP kinases, on appressorium formation during the last decade. More recently, pharmacological data suggest that calcium/calmodulin-dependent signaling system is involved in its appressorium formation. To determine the role of phosphoinositide-specific phospholipase C (PI-PLC) on appressorium formation, a gene (WPLCl) encoding PI-PLC was cloned and characterized from M. grisea strain 70-15. Sequence analysis showed that MPLCl has alt five conserved domains present in other phospholipase C genes from several filamentous fungi and mammals. Null mutants (mplcl) generated by targeted gene disruption exhibited pleiotropic effects on conidial morphology, appressorium formation, fertility and pathogenicity. mplcl mutants developed nonfunctional appressoria and are also defective in infectious growth in host tissues. Defects in appressorium formation and pathogenicity in mplcl mutants were complemented by a mouse PLCdelta-1 cDNA under the control of the MPLCl promoter. These results suggest that cellular signaling mediated by MPLCl plays crucial and diverse roles in development and pathogenicity of M. grisea, and functional conservation between fungal and mammalian Pl-PLCs.

Magnaporthee grisea causes the devastating blast disease of rice. Entensive research has been conducted on infection mechanisms, particularly on appressorium formation and penetration, of this fungus during the last decade. However, the role(s) of cell-wall-degrading enzymes (CWDEs) on pathogenesis is not clearly demonstrated at molecular level. Many CWDES in plant pathogenic fungi including M. grisea are redundant; that is, there are multiple genes encoding enzymes with a similar or overlapping spectrum of activities. It is laborious to isolate all of the genes encoding related enzymes and to construct mutants lacking all 9f them. Thus, we considered alternative strategies to address the role of CWDEs in pathogenesis. Since expression of CWDE genes Is repressed by a simple sugar, as the first step, we cloned a Snfl (sucrose non-fermenting) gene (MgSnf1) from M. grisea. The predicted amino acid sequence showed a high identity with other Snf1 genes from various fungi. To elucidate molecular function of MgSnf1, a transformant lacking MgSnf1 was created by targeted gene replacement. En glucose, sucrose, and xylan the MgSnf1 mutant grew normally but in pectin and complex media, it grew slower than wild type. Expression of various CWDEs in MgSnf1 mutant was investigated and found that expression of some CWDEs is repressed. However, no significant difference was observed in conidial germination, appressorium formation, and pathogenicity in MgSnf1 mutant. However, MgSnf1 functionally complemented a yeast MgSnf1 mutant. These results suggest that MgSnf1 is involved in regulation of CWDEs and MgSnf1 is dispensable in pathogenicity of M. grisea.

In most ascomycetes, a single mating type locus, MAT, with two alternate forms (MAT1-1 and MAT1-2) called idiomorphs, controls mating ability. In heterothallic ascomycetes these alternate idiomorphs reside in different nuclei. In contrast, most homothallic ascomycetes carry both MAT1-1 and MAT1-2 in a single nucleus, usually closely linked. An example of the latter is Gibberella zeae, a producer of mycotoxins such as trichothecene and zearalenone that threaten human and animal health. We asked if G. zeae could be made strictly heterothallic by manipulation of MAT. Targeted gene replacement was used to differentially delete MAT1-1 or MAT1-2 from a wild type haploid MAT1-1 MAT1-2 strain, resulting in MAT1-1；mat1-2, mat1-1；MAT1-2 strains that were self-sterile, yet able to cross to wild type testers and more importantly, to each other. These results indicated that differential deletion of MAT idiomorphs eliminates selfing ability of G. zeae, but the ability to outcross is retained. To our knowledge, this is the first report of complete conversion of fungal reproductive strategy from homothallic to heterothallic by targeted manipulation of MAT. Practically, this approach opens the door to simple and efficient procedures for obtaining sexual recombinants of G. zeae that will be useful for genetic analyses of mycotoxin production and other traits, such as ability to cause disease.

Fusarium graminearum is an important pathogen of cereal crops in many areas of the world causing head blight and ear rot of small grains. In addition to serious economic losses, this fungus produces mycotoxins, such as trichothecenes and zearalenone on diseased crops and has been a potential threat to human and animal health. To massively identify pathogenesis-related genes from F. graminearum, two representative strains (SCKO4 from rice and Z03643 from wheat) were mutagenized using restriction enzyme-mediated integration (REMI). In total, 20,DOD REMI transformants have been collected from the two strains. So far, 63 mutants for several traits involved in disease development such as virulence, mycotoxin production, and sporulation have been selected from 3,000 REMI transformants. Now, selected mutants of interest have being genetically analyzed using a newly developed outcross method (See Jungkwan Lee et al poster). In addition, cloning and characterization of genomic DNA regions flanking the insertional site in the genome of the mutants are in progress.

Powdery mildew on strawberry plants, caused by Spherotheca aphanis (Wallr.) U. Braun var. aphanis, is the most serious disease for strawberry production. There is a demand to develop the substitutes for chemicals which are more environment friendly materials. Control of powdery mildew was evaluated on Akihime, Sachinoka, Dochiodome, Noyho and Redpearl varieties grown in the greenhouse. Applications of 1％, 0.5％, and 0.25％ of sodium bicarbonate (NaHCO3Nyoho in greenhouse experiments. Non-phytotoxicity was revealed on the leaves and fruits of strawberry at these concentrations. This$.$result indicates that a mixture of sodium bicarbonate and tween 20 is a useful substitute for fungicides to control powdery mildew of strawberry.

Long-term repeated culturing of biocontrol agents on a certain medium often results in reduced biocontrol efficacy and altered physiology. Effect of culturing media on biocontrol ability and physiological state of Burkholderia gladioli strain B543 was investigated. Over 20 times repeated cultivation of B. giadioli strain B543 on Kings B medium or nutrient agar medium showed improved biological control of cucumber damping-off caused by Pythium ultimum, while one time cultivation on KB or NA did not. The repeated cultivation also induced the physiological changes of the biocontrol agent such as antifungal activity and the production of protease and siderophore. Our result indicates that adaptation to proper culturing medium can alter biocontrol ability and must consider in optimizing the use of biocontrol agents.

TiO$_2$photocatalytic ozonation was attempted to disinfect fungal pathogens causing postharvest spoilage of kiwifruits and to decompose fungicide residuals on kiwifruits. TiO$_2$Photocatalytic ozonation process synergistically degraded organic compound and inhibited conidial germination of the fungal pathogen compared to single treatment of ozonation or photocatalysis. The efficient control of fungal spoilage and degradation of residual fungicide on kiwifruits indicate that TiO$_2$photocatalytic ozonation is a very attractive method for postharvest disease control of kiwifruits as an alternative to fungicides application.

Antogonistic bacteria against Rhizoctonia solani and Pythium spp., causing serious damage to golf course grasses, were isolated from the top soil of several golf courses in Korea. The isolate of Limk0102 was selected as the biological agent by characterization of antifungal activity, large scale preparation, fungicides tolerance and ecological fitness to the targe environment. The isolate was identified as Bacillus subtilis by biochemical and physiological characterization, and 165 rDNA sequence analysis. The bacterial agent was formulated as a granule type by seeding it on fine sand. The formulated agent showed high recovery rate (more than 10$\^$8/ cells/g sand) even after 6 month-storage at room temperature with similar antifungal activity with that of original cells. In vitro, the biological agent successfully exhibited antagonistic performance on bentgrass inoculated with R. solani or Pythium spp. isolated from the diseased grasses on golf courses. Field evaluation on disease control activity and ecological fitness of the agent is now under going on several golf courses.

Changes of control efficacy of metalaxyl to potato late blight caused by Phytophthora infestans were examined in potato fields from 2001 to 2003. In 2001 and 2002, control efficacy of metalaxyl was similar to those of dimethomorph and ethaboxam. However, the control efficacy of metalaxyl were decreased to 50.3％ in 2003. Total 366 isolates of P infestans obtained from several areas in Korea from 2001 to 2003 were examined for changes of sensitivity to metalaxyl. About 6.8％ of fungal isolates examined in 2001 were sensitive, 84.1％ were intermediate resistant, 9.1％ were resistant to metalaxyl. Among the isolates collected in 2002, 3.9％ were sensitive, 75.6％ were intermediate resistant, 20.6％ were resistant to the chemical. However, among the isolates obtained in 2003, 55.9％ were intermediate resistant, 44.1％ were resistant, but none of the isolates tested were sensitive. Both A1 and A2 mating type isolate were isolated in 2002∼2003. However, all isolates collected in 2001 were A1 mating type. About 87.5％ of the isolates collected in 2002, 89.8％ In 2003 were determined as A1 mating type. The majority of the p. infestans isolates were A1 mating types. Changes of control efficacy of metalaxyl to potato late blight might be caused by the occurrence rate of moderately resistant isolates within A1 mating type.

Disease suppression by ectomycorrhizal(ECM) fungi has been demonstrated on red pine seedlings. Culturing of pathogenic fungi on petri plates containing culture filtrates of ECM fungi showed that culture filtrates of the ECM fungus Hebeloma cylindrosporum may inhibit the mycelial growth of all tested soil-borne plant pathogenic(SBPP) fungi upto 60％, In order to examine the effects of ECM fungi on SBPP fungi and on red pine seedlings, both symbiotic and pathogenic fungi were inoculated into the soil with red pine seedlings by three inoculation methods; pre-inoculation of SBPP fungi 10 days before inoculation of ECM fungi, simultaneous inoculation of both fungi, post-inoculation of SBPP fungi 60 days after inoculation of ECM fungi. Seedling mortality, seedling growth, and ectomycorrhizal formation by the combined treatments were examined and compared. Pine seedlings were dead by the pre-inoculation of pathogenic fungi, except Rhizina undulate which required 9-12 days, within 6 days after inoculation. Among pathogenic fungi tested, Fusarium oxysporum was the most pathogenic with the mortality of 44％. However, no dead seedlings were shown by simultaneous inoculation of both fungi or pre-inoculation of ECM fungi. In addition, pine seedlings treated by simultaneous or post-inoculation of SBPP fungi were relatively higher than those treated by pre-inoculation in diameter at root crown and the number of ectomycorrhizal roots. There were no significant differences among inoculation methods in root length and dry weight of treated seedlings. It means that ECM fungi somehow play a role in protecting primary roots of red pine seedlings against invasion by the SBPP fungi.

Potato late blight, caused by Phytophthora infestans, is one of the important diseases in potato cultivation areas. Though the incidence of late blight was depend on the inoculums and climatic condition In each fields, the foliar blight was reached to 100％ under the severe disease pressure condition in 2003. Outbreak of foliar blight was concentrated from May and July and evaluation of ten fungicides to control of late blight was made at Daekwallryoung area in potato fields. Based on the company recommendation, those fungicides were applied by a sprayer at the recommended rates on a weekly application schedule. Effect of ten fungicides on foliar blight was based on area under disease progress curve (AUDPC). Across all fungicides was reduced by 77％ in AUDPC and dimethomorph was reduced by 92％ in AUDPC during the same period, respectively. Those fungicide were inhibited the mycelial growth of isolate with different rate in chemical amended medium and several fungicides were completely limited the growth of isolate.

Colonization of Pseudomonas chlororaphis O6, a nonpathogenic rhizobacterium, on the roots induced systemic resistance in cucumber plants against tai-get leaf spot, a foliar disease caused by Corynespora cassiicola. A cDNA library was constructed using mRNA extracted from the cucumber leaves 12 h after inoculation with C. cassiicola, which roots had been previously treated with O6. To identify the genes involved in the O6-mediated induced systemic resistance (ISR), we employed a subtractive hybridization method using mRNAs extracted from C cassiicola-inoculated cucumber leaves with and without previous O6 treatment on the plant roots. Differential screening of the cDNA library led to the isolation of 5 distinct genesencoding a GTP-binding protein, a putative senescence-associated protein, a galactinol synthase, a hypersensitive-induced reaction protein, and a putative aquaporin. Expressions of these genes are not induced by O6 colonization alone. Before challenge inoculation, no increase in the gene transcriptions could be detected in previously O6-treated and untreated plants but, upon subsequent inoculation with the pathogenic fungus, transcription levels in O6-treated plants rose significantly faster and stronger than in untreated plants. Therefore, the O6-mediated ISR may be associated with an enhanced capacity for the rapid and effective activation of cellular defense responses which becomes apparent only after challenge inoculation on the distal, untreated plant parts, as suggested by Conrath et al. (2002). This work was supported by a grant R11-2001-092-02006-0 from the Korea Science and Engineering Foundation through the Agricultural Plant Stress Research Center at Chonnam National University.

In commercial greenhouses, senescent flower petals or flowers of vegetables such as tomato, strawberry, hot pepper and zucchini squash were blighted to be removed from fruits within five days after spraying of Trichoderma harzianum YC459 (TORY), a biocontrol agent for the gray mold rot of vegetables caused by B. cinerea The mechanism for selective colonization of senescent floral tissues by T. harzianum YC459 was elucidated using fresh and senescent (Hays and 14days after flowering, respectively) floral tissues of zucchini squash (Cucurbita moschata Duchesne). The spores of T. hrzianum YC459 were produced more on agar and liquid culture media supplemented with 5％ dry powder of senescent floral tissues than fresh tissues during 15days. Mycelial growth was also much better in the media with senescent tissues than with fresh tissues. Enzyme activities of amylase, polygalacturonase and cellulase in the liquid media which might be involved in the colonization of tissues by T. harzianum YC459 were compared. The activities of three enzymes were much higher in the media with senescent floral tissues than with fresh floral tissues reaching to the maximum during 9 to 12days of incubation. Based on the results, the removal of senescent floral tissues, a possible inoculum source of the pathogen, may be another mechanism for biocontrol of gray mold rot of vegetables by T. harzianum YC459.

Bakanae disease, caused by Gibberella fujikuroi (anamorph Fusarium moniliforme J. Sheldon), a typical seed-borne disease of rice occurs from nursery to paddy fields. Consequently, chemical seed disinfectants is the most efficient control method. Several seed treatment methods with various fungicides were attempted to inhibit disease. Spray and 24 hrs immersion of seeds using prochloraz emulsion reduced disease infection and the control value were 99.3 and 100％, respectively. In contrast, dressing to wet seeds thiophanate-methyl＋thiram wp and benomyl＋thiram wp reduced disease infection more effectively than 24 hrs immersion of seeds. However, dressing of carpropamid+imidacloprid＋fludioxonil wp to wet seeds did not reduced disease as well as wettable liquid of fludioxonil. The results suggest that the bakanae disease might be disinfected effectively by 24 hrs immersion of seeds in prochloraz emulsion and seed dressing of fungicides.

In order to elucidate influence of nutritional status on rice brown spot caused by Cochliobolus miyabeanus, rice cultivation soils and rice straws were collected from paddy fields where ice brown spot occurred severely, moderately, a little and none respectively. Rice plant materials were analyzed to measure inorganic nutrients in rice straws and rice seeds. Analysis of chemical properties of rice paddy soil showed that EC and contents of available phosphate, cation and silicic acid in soil with severe infections were lower than those in healthy soil. This result suggests that amount and holding capacity of nutrient contents in soils collected from paddy field with infection of C. miyabeanus are relatively low compared to those in soils collected from healthy paddy field. Analysis of inorganic nutrients in rice straws showed that amount of macronutrient elements such as silicic acids, available phosphate and total nitrogen, and micronutrients such as copper, iron and zinc in rice straws from paddy field with infection were lower than those in healthy soil. Especially amount of iron and silicic acid were very low in rice straws from paddy field soils with infection Amount of inorganic nutrients such as iron and zinc in rice seeds was the same trend as those of rice straws. These results showed that one of major factors affecting rice brown spot was amount of nutrient contents in soil and rice straw.

In order to develop an effective spray program for control of apple white rot with reduced use of fungicides, the control efficacy of several fungicides that has been intensively used for control of the disease was assessed. They were sprayed on the same tree with 15 day interval from late May to early August. Just prior to and after each spray, 100 fruits were bagged with two layered fruit bag to limit the chemical application in only one time, and the disease incidence and latent infection frequency on the bagged apples were examined. Some fungicides such as folpet, iminoctadine-triacetate and azoxystrobin showed a high post-infectional activity even though the former two are non- systemic. Folpet suppressed symptom development, iminoctadine-triacetate reduced infection frequency and azoxystrobin acted in both ways. When those fungicides were ！！ adopted in a spray program, once in a cropping season, their post-infectional activity became much greater. This activity shown by the non-systemic fungicides was supposed to be derived from the peculiar infection process of the white rot fungus of which the pathogen is usually remain latent in the corked cells of lenticel until the apple reach mature stage.

Severe soybean-sprout rot was found at the mass productive factory in 2000 and 2001 and it caused 10-20％ loss of the production. Pythium sp. was isolated almost 90％ by potato dextrose agar from rotted root and hypocotylsof the sprouts. And the pathogencity tests using test tubes with 2％ water agar and small containers (30 ${\times}$ 30 ${\times}$ 50 cm, WxLxH) cultivation were shown a similar rot on roots and hypocotyls. The fungal mycelium grew rapidly on the water agar and it prevented the seed germination. Density of the Pythium sp. in the recycled water system at the factory was periodically measured using a selective medium, corn meal agar with Pimaricin 10 mg, Rifampicin 10 mg, Ampicillin 100 mg per 1 liter in order to check the contamination of recycled water. After fitering step using 5 and 1 ml in the recycled system was applied and it was effectively controlled Pythium rot. The daily yield of sprout was stable and the occurrenceof Pythium in the recycled water was much less after filtering. The fungal isolates were identified as Pythium deliense Meurs based on various mycological characteristics on corn meal agar and sucrose-asparagine bentgrass leaf culture medium. P. deliens oogonia were spherical, smooth, 19-23 urn in diameter, and their stalk bending toward antheridia. Antheridia were straw hat-shaped, curred club-shaped, therminal or intercalary, monoclinous, occasionally diclinous, 12∼15 ${\times}$ 8∼11 um, 1(∼2) per oogonium.

A growth chamber fumigation was conducted to evaluate the ozone (O3) on the physiology of three hot pepper, Capsicum annuum L. cultivars, 'dabotab', 'buchon' and 'pochungchun'. Thirty-day old plants were exposed to O3 of 120 nl 1-1 in the chambers for 8 h d-1 for 3 days. Foliar damage due to O3 was different from the varieties, 'dabotab'was most sensitive to O3, 'pochungchun' was medium, and 'buchon' was resistant. Ozone symptom on the leaves was bifacial necorsis. Photosynthesis and stomatal conductance were decreased due to O3 treatment, but they were not much different from the variety. Decreases of net photosynthesis by O3 were 56％, 40％ and 35％ on 'dabotab', 'buchon' and 'pochungchun', respectively Decreases of stomatal conductance by O3 were 66％, 63％, and 50％ on each varieties. Ozone closed the stomata and decrease net photosynthesis on hot peppers regardless of the variety. Light curves on the three varieties were showing similar patterns that O3 damage on net photosynthesis were started at the low levels of light with or without the visible injury, Assimilation-internal CO2 concentration curves of the three cultivars were not different due to the treatment. It means there was not significant biochemical damage Inside the leaves by O3. In conclusion, ozone closed the stomata and damaged light capturing system of the pepper leaves with or without the visible damage. Although visible damage of the leaves could be a good indicator of O3 resistance, the ecophysiological change by O3 were not proportional to the amout of visible injuries

In our previous studies, we selected three antagonistic bacteria, KJ1R5, KJ2C12, and KJ9C8 against Phytophthora capsici, the casual agent of Phytophthora blight of pepper. For elucidating production, root colonization, and total microbial activity were investigated. The dual culture assay was accomplished to elucidate existence of antibiotics. In this assay, any antagonistic bacteria did not inhibit growth of six important fungal plant pathogens, suggesting that these antagonists do not produce antibiotics. root surface or rhizosphere soil colonizations were examined with spontaneous rifampicin-resistant mutants equal to antagonistic ability of wild types. KJ2C12 colonized consistently rhizosphere soil while yellowish colonies of KJ1R5 and KJ9C8 well colonized root surfaces and rhizosphere soil. Total microbial activity in pots treated with the antagonistic bacteria was measured using fluorescein diacetate hydrolysis. total microbial activity of three antagonistic bacteria treatments was significantly higher than that of buffer-treated control until 4days after treatment. However, total microbial activity of treatment of three antagonistic bacteria decreased after 7 days. These results indicate that the antagonistic bacteria, KJ1R5 and KJ9C8 colonized and protected roots well against Phytophthora blight of pepper through competition of infection courts, especially competitions.

Control of bacterial blossom blight of kiwifruit (Actinidia deliciosa) has been mainly depended on chemical control. Recently cultural practices such as trunk girdling of kiwifruit trees and rainproof installation over kiwifruit trees also were conducted as the alternative control practices. Each of the control methods was evaluated for the best practicing conditions for the control of bacterial blossom blight of kiwifruit. Among the various combinations of spray times and spray periods, optimum spray program of antibiotics was turned out to be 3 times with intervals of 10 days from early May during the flowering season of kiwifruits. Optimum periods of trunk girdling of kiwifruit trees were from late March to late April. Trunk girdling with 20-30 mm wide showed best control efficacies on bacterial blossom blight, irrespective of the heights of girdling on trunks of kiwifruit trees. Optimum period of rainproof installation over kiwifruit trees was from March till late April, irrespective of installation methods.

E. intermedium 60-2G possessing a strong ability to solubilize insoluble phosphate, has plant growth-promoting activity, induced systemic resistance activity against scab pathogen in cucumber, and antifungal activity against various phytopathogenic fungi. The phosphate solubilizing activity of 60-2G may be mainly accomplished by production of gluconic acid through a direct extracellular oxidation of glucose by glucose dehydrogenase that required a PQQ cofactor for its activation. A pqq gene cluster conferred Phosphate-solubilizing activity in E. coli DH5${\alpha}$ was cloned and sequenced. The 6,783 bP pqq sequence had six open reading frames (from A to F) and showed 50-95％ homology to pqq genes from other bacteria. The E. coli strain expressing the pqq genes solubilized phosphate from hydroxyapatite after a pH drop to 4.0, which paralleled in time the secretion of gluconic acid. To study the role of PQQ in biocontrol traits of E. intermedium, PQQ mutants of 60-2G were constructed by marker exchangee mutagenesis. The PQQ mutants of E. intermedium were lost activities of solubilizing phosphate, growth inhibition of phytopathogenic fungi, and plant growth promotion. These findings suggest that PQQ plays an important role, possibly activation of certain enzymes, in several beneficial bacterial traits of E. intermedium by as yet an unknown mechanism.

Plant growth promoting rhizobacteria (PGPR) have been shown to suppress phytopthora blight. This suppression has been related to both microbial antagonism and induced resistance. The PGPR isolates were screened by dual culture plate method and most of the isolates were showed varying levels of antagonism. Among the PGPR isolates pyoverdin, pyochelin and salicylic acid producing strains showed the maximum inhibition of mycelial growth of Phytopkhora capsici and increased plant growth promotion in red pepper. PGPR isolates further analysed for its ability to induce production of defence related enzymes and chemicals. The activities such as Phenyle alanin ammonia Iyase (PAL), Peroxidase (PO), Polyphenol oxidase (PPO) and accumulation of phenolics were observed in PGPR pretreated red pepper plants challenged with Phytopkhora capsici. The present study shows that an addition of direct antagonism and plant growth promotion, induction of defense related enzymes involved to enhance resistance against invasion of P. capsici in red pepper.

In order to develop a new microbial fungicide for the control of vegetable diseases using endophytic bacteria, a total of 260 bacterial strains were isolated from fresh tissues of 5 plant species. After they were cultured in broth media, their antifungal activities were screened by in vivo bioassays against Botrytis cinerea(tomato gray mold), Pythium ultimum(cucumber damping-off), Phytopkhora infestans(tomato late blight), Colletotrichum orbiculare(cucumber anthracnose), and Blumeria graminis f. sp. hordei(barley powdery mildew). As the results of screening, 38 bacterial strains showed potent antifungal activities against at least one of 5 plant pathogens. A bacterial strain EB072 displayed potent disease control activities against 3 plant diseases. Among the bacterial strains with a potent antifungal activity against cucunlber anthracnose, three bacterial strains, EB054, EB151 and EB215, also displayed a potent in vitro antifungal activity against C. acutatum, a fungal agent causing pepper anthracnose. A bacterial strain EB215 obtained from roots of cucumber was identified as Burkholderia cepacia based on its physiological and biochemical characteristics and 165 rRNA gene sequence. An antifungal substance was isolated from the liquid cultures of B. cepacia EB215 strain by ethyl acetate partitioning, repeated silica gel column chromatography, and invitro bioassay, Its structural determination is in progress by various instrumental analyses.

Serratia plymuthim A21-4 strongly inhibits the mycelial growth, zoospore formation, and cystospore germination of Phytophthor spp and Pythium species. The bacterial isolate produced antifungal substance and chitinase. The bacteria also enhanced to plant growth remarkably in low nutritional condition. The application of cell suspension of A21-4 to pepper seedlings in greenhouse experiments and soil drenching in farmer's field was proved successfully to control the phythophthora blight of pepper. For the effective control, however, relatively high density of cell number(10$\^$9/cfu/$m\ell$) is required. Density effect was similar in plant growth promoting activity of A21-4. Though this investigation we improved the problem with changes of culture condition of bacteria and some nutritional amendment.

More than 1200 microorganisms were isolated from soil samples collected from various sources and localities. Among the isolates, 2 actinomyces (TH-04 and BA313) and 1 Bacillus sp. (CJ3) were selected as antagonists to pepper anthracnose fungus Colletotrichum gloeosporioides. These 3 isolates inhibitied mycelial growth of C gloeosporioides and the inhibition rates were over 70％ on PDA. When the isolates were co-cultured with conidia of C. gloeosporioides in potato dextrose broth, conidial germination was severely inhibited and the inhibition rates of TH-04, BA313, and CJ3 at 24 hours were 75％, 72％, and 68％, respectively. The inhibition rates at n hours incubation were not much different from the rates at 24 hours. To check the activity on the plant, each isolate was mixed with equal volume of conidial suspension of C. gloeosporioides and wound-inoculated on green pepper fruit. After 6 days, the anthracnose lesions on the fruits inoculated with the mixture were much smaller than the lesions caused by the C. gloeosporioides itself. The lesion areas of TH-04 or BA313 treated pepper were less than 30％ of the check. TH-04 and BA313 also showed antagonistic activity to Phytophthora spp. and Botrytis cinerea. By scanning electron microscopy and fatty acid analyses (MIDI), TH-04 and BA313 were identified to Streptomyces halstedii and S. violaceusniger, repectively.

An antagonistic bacterium, KJ1R5,, to Phytophthora capsici was obtained from root interior of a healthy pepper plant. To identify the bacterial antagonist, 16S rDNA sequence analysis, Biolog system, fatty acid methyl-esters (FAMEs), and physiological and biochemical characterization were conducted. The determined 165 rDNA sequence of KJ1R5, showed higher similarities to those of a group consisting of several Chryseobacterium strains with 95.2, 95.2, and 95,1％ similarity to C. defluvii, Chryseobacterium sp. FR2, and C. scophthalmum, respectively, In addition, Halounella gailinarum, Bergeyella zoohelcum, and Riemerella anatipestifer are another group for KJ1R5, with 94.1, 89.7, and 87.2％ similarities, respectively When identification of the antagonistic bacterium, KJ1R5, was conducted using BIOLOG system, the strain KJ1R5, was identified as Flavobacterium tirrenicum (similarity； 0.75％). Fatty acid profiles of the strain KJ1R5, were composed mainly of iso-17：0 w9c and iso-15：0 and identified as Chryseobacterium balustinum (similarity 0.524％). KJ1R5, was Gram-negative, regular short rods ranging from 0.8 $\mu\textrm{m}$ to 1.0 $\mu\textrm{m}$ and had no flagella. Phenotypic characterization of the antagonistic bacterium indicated that KJ1R5, were included in the genus Chreseobacterium, which belongs to the family Flavobacteriaceae. The strain was distinguished from these six existing species. These results indicated that strain might be placed as a new species in the genus Chryseobacterium.

A new and effective formulation using antagonistic bacteria, Burkholderia cepacia YC5025 in vegetable oil was developed for the biocontrol of anthracnose. The bacterial population in the formulation was maintained to 5x10/sup7/ cfu/ml upto 60 days at room temperature. Control efficacy of the formulation for anthracnose was over 80％ by spraying of diluted suspension(x1,000) in growth chamber tests. On the contrary, the bacterial suspension in distilled water or bacterial culture broth containing same number of spores as the formulation had low control efficacy around 40％ even 2-weeks storage after preparation. The shelf-life of the formulation was longer than that of bacterial preparation using clay minerals such as talc or bentonite. The mechanisms of newly developed bacterial formulation are possibly the formation of water film on the surface of cucumber leaves and inactivation of the bacteria in the vegetable oils during storage. Further field tests and improvements with new liquid bacteiral formulation need to be done for practical application.

Antagonistic effect of bacterial strains isolated from phylloplane of strawberry plants grown In greenhouse was tested on Botrytis cinerea Among the promising bacterial strains, Bacillus sp. S1-0210 showed highest inhibition of mycelial growth of B. cinerea and a broad spectrum of antifungal activities against many plant pathogenic fungi in vitro. Bacillus sp. S1-0210 was identified as Bacillus subtilis based on the analysis of 185 rDNA as well as its biochemical characteristics. Application of wettable powder formulation of B. subtiiis S1-0210 significantly reduced the incidence of gray mold on trawberry fruits during storage. Results showed that treatment of B. subtilis S1-0210 decreased the incidence of gray mold by 4.8％ whereas the incidence in control was 77.9％, indicating that the formulation of B. subtilis S1-0210 will be practically applied on strawberry fruits as a biocontrol agent of gray mold during storage.

Bacterial canker of sweet cherry (Prunus cerasus L.) was observed in farmers' orchard in Goesan, Chungbuk in 2003. Typical canker symptom occurred on the branches or twigs of sweet cherry in early spring and bacterial exudates oozed out of the cracked barks of diseased trees. Watersoaked brown symptom appeared on the leaves and severe infection caused thorough defoliation on the branches or twigs of sweet cherry. When cut the severely infected branches or twigs, irregular and rusty-colored symptoms in sapwood and heartwood were clearly found, indicating that they could serve as specific symptoms of bacterial canker of sweet cherry. The gram negative, aerobic bacterium isolated from the lesion produced fluorescent pigments on King's B agar medium but did not grow at 37$^{\circ}C$ The bacterium formed Levan-type colonies, and showed negative reactions in oxidase reaction, arginine dihydrolysis test, and pectolytic activity Based on the biochemical and pathological characteristics, the causal organism was identified as Pseudomonas syringae pv. morsprunorum. This is the first report on bacterial canker of sweet cherry in Korea.

E. intermedium Blocontrol activity of a P. chlororaphis rhizobacteium O6, depends to the synthesis of extracellular secondary metabolites and exoenzymes, thought to antagonize the pathogenicity of a variety of phytopathogenic fungi. The production of secondary metabolites and exoenzymes in O6, depends essentially on the GacS-mediated signal transduction pathway, which activates largely unknown signal transduction pathway. To exploit the GacS-mediated signal transdcution pathway involved in activation of ph genes that are necessary for biosynthesis of phenazine from P. chlororaphis O6, we cloned and sequenced the phz operon, rpoS gene encoding stationary specific sigma factor, ppx gene encoding polyphosphatase, and lon gene encoding ion protease. Expression of each gene in wild type and GacS mutant were analyzed by RT-PCR. Transcripts from rpoS, phzI enconing acylhomoserine lactone (AHL) synthase, and ph structural genes in the GacS mutant were reduced in each of these growth phases compared to the wild type. The GacS or Lon mutant was found to be deficient in the production of phenzines, exoenzymes, and the acylhomoserine lactone. These mutants were not complemented by ph operon and addition of exogenous AHL. These results indicate that the GacS global regulatory systems controls phenazine production at multiple levels. Future research will focus to identifying the GacS-mediated regulatory cascade involving in production of phenazine in P. chlororaphis.

An attempt was made to screen antifungal activities of Bacillus thuringiensis strains on various plant pathogens, Botryosphaeria dothidea, Diaporthe actinidiae, Botrytis cinerea, Glomerella cingulata, Colletorichum cocodes, Sclerotinia scierotiorum, Alternaria alternata, Helicobuidium mompa, Bipolaris coicis, Fusarium graminearum and Rhizoctosnia solani. Ten and forty-five strains of B. thuringiensis were isolated from animal feces in Korea and Japan, respectively. Inhibitory effects of the strains on the mycelial growth of the pathogens were examined on the mixed media of potato dextrose agar and nutrient agar. Approximately half of the strains inhibited the mycelial growth of one or more pathogens. Most of the pathogens were inhibited by any of the strains but Fusarium graminearum and Rhizoctonia solani were not inhibited at all. This is the first report that B. thuringiensis shows a potent antifungal activity on plant pathogens in Korea.

A dctA gene encoding a protein with identity to a C4-dicarboxylate/H+ was cloned from a beneficial biocontrol bacterium, P. chororaphis O6. Expression of the dctA was induced in minimal medium by several organic acids and was repressed by glucose. Highest expression was observed in early-log cells grown on fumarate and succinate with decline as cells approached late-log phase. The dctA transcript accumulated weakly when cells were grown on malate but strong expression was observed with benzoate. Expression of the dctA transcript was repressed in early-log cells upon addition of glucose to fumarate, but was detected as the cell culture aged. A dctA-deficient mutant of O6, constructed by marker exchange mutagenesis, did not grow on minimal medium containing succinate, benzoate, or fumarate, and growth on malate was delayed. The dctA mutant and wild type grew equally on glucose. The dctA mutant on cucumber roots in sterilized potting soil was colonized at levels comparable to those of the wild type, but induction level of disease resistance by the mutant against target leaf spot disease was decreased. These results may indicate that the dctA is essential for utilization of certain organic acids and its expression is controlled by the availability of sugars. In addition, the dctA is not essenitial for cucumber root colonization, but important for induction of disease resistance.

The global regulator, GacS (global antibiotic and cyanide sensor kinase), was required for the increased resistance to hydrogen peroxide occurring as cultures of the rhizobacterium, P. chlororaphis O6, matured. Specific stationary-phase peroxidase and catalase isozymes were absent in the GacS mutant, whereas a manganese-superoxide dismutase isozyme was expressed earlier and to a great extent than wild type. In the wild type cell, transcript accumulation of rpoS was higher in late logarithmic-phase cells than cells from mid logarithmic- or stationary-phase. Transcripts from rpoS in the GacS mutant were reduced in each of these growth phases compared to the wild type expression. The down stream sequence from rpoS lacked sequences encoding a small RNA, rsmZ, found in other pseudomonads and implicated in control of genes activated by the GacS system. These findings suggest that GacS-mediated regulation of RpoS plays role in control of oxidative stress in P. chlororaphis O6 by as yet an unknown mechanism.

Lipopolysaccharide, siderophore, and cyclic dipeptide have been shown to be necessary for ISR induction by pseudomnads. However, there is no report on cloning of genes or generating specific mutants involving in ISR activity. A biological control bacteium P. chlororaphis O6 induces resistance to Erwinia carotovora subsp. carotovara SCCI in tobacco and induces drought resistance in Arabidopsis. To isolate genes involved in ISR activity and induction of drough resistance of O6, we constructed Tn5 mutants and were used to screen for ISR activity and drought resistance activity using microtiter assay with tobacco and Arabidopsis. Thirty-three ISR-deficient mutants were selected, and the nine ISR-deficient mutants were also lost activity of drought resistance. The flanking sequence analysis of the ISR and drought resistance-deficient mutants showed that a gacS gene encoding a two-component sensor kinase, and a mce gene encoding a protein involved in mycobacterial cell entry were mutated. The flanking sequence of each Tn5 mutant altered ISR activity is currently under investigation. These results indicate that gacS and mce are important genes in induction of ISR activity and drought resistance of P. chlororaphis O6. Our works will open opportunities for identification of bacterial genes or traits that are involved in ISR activity and induced drought resistance of P. chlororaphis O6.

The ability of P. chlororaphis O6 to induce resistance to Erwinia carotovora subsp. carotovara SCCI and to promote growth in tobacco was demonstrated in microtiter assays on plants pre-inoculated at the root level with the bacteria before challenge with the leaf pathogen. To identify th bacterial determinants involved in induced systemic resistance and plant growth promotion, cell culture of O6 grown in King's medium B was fractionated with organic solvents and purified using various columns. in vivo and in vitro assays with samples from successive fractionation steps of the O6 supernatant led to the conclusion that antibacterial compounds were observed in aqueous layer, and to the isolation of fractions containing metabolites that retained most of the resistance-inducing activity (70：30, methanol：water) and the plant growth promotion (80：20 and 90：10, methanol：water) after ODS column chromatography. Although these molecules remain to be purified further and structurally characterized, its isolation is an addition to the range of determinants from plant growth-promoting rhizobacteria known to stimulate plant defence.

A bacterial strain YC4963 with antifungal activity against Colletotrichum orbiculare, a causal organism of cucumber anthracnose was isolated from the rhizosphere soil of Siegesbeckia pubescens (Siegesbeckia pubescens Makino；Family：Compositae) in Korea. Based on physiological and biochemical characteristics and 16S ribosomal DNA sequence analysis, the bacterial strain was identified as Pseudomonu aureofaciens. The bacteria also inhibited mycelial growth of several plant fungal pathogens such as Botrytis cinerea, Fusarium oxysporum and Rhizoctonia solani on PDA and 0.1 TSA media. The antibiotic activity was found from the culture filtrate of TSB(tryptic soy broth) and its active compounds were quantitatively bound to XAD adsorber resin. The antibiotic spectrum was broad and growth of C. orbiculare and F. oxysporum, B. cinerea were inhibited at very low concentration. The chemical data from various chromatographic procedures showed that active fraction consisted of at least two phenazine derivatives. However, the metabolites had no inhibitory effect on Pythium ultimum which was reported to be sensitive to phenazine antibiotics. The compounds responsible for the activity are now under investigation.

This studies were investigated the occurrence of Sclerotinia rot by Sclerotinia Sclerotiorum at the lettuce field in Uiryeong-Gun, Gyeongsangnam-Do and were isolated the most effective microorganism for the biological control to the pathogen, S. sclerotiorum YR-1 strain from diseased soil and lettuce leaves. For the pathogenicity test, the most suitable inoculmn density of YR-1 strain was selected as the mycelial suspension of 40m1 showing disease incidence of 80％, and the symptom showed as same as at the fields, the leaves and stem had rotten and developed white downy mycelial at the diseased lesion on the leaves and stems, and produced black and irregular sclerotinia. On the PDA dual test, about 300 isolates were examined the antifungal activity to the pathogen, YR-1 strain, and among them, A-2, A-7, and RH-4 strain were selected most effective antagonistic bacteria. At pots test, the control value of A-7 strain showed the highest value as 85％ which was more effective than that of others in a growth chamber. For the promotion of control effect, the selected 3 isolates were spayed on the lettuce leaves as a sole and/or mixed treatments in a growth chamber, the mixed treatment of A-7 and RH-4 strain showing the control value of 90％ was most effective than that of sole treatment with A-7 or RH-4 strain showing the control value of 80％, respectively and mixed treatment with A-2 and A-7 strain and A-2, A-7 and RH-4 strain. In addition, 3 bacteria re-isolated from diseased soils, and all of the selected 6 isolates investigated the control effect at pots in a growth chamber, According to the results, A-7 and Pro-EB 15 strain showed the control value of 91.0％ and 90.1％ respectively, and they were selected most effectual antagonistic bacteria to control lettuce sclerotinia rot and identified as the Bacillus mojanuinensis by 16s RNA analysis. This is the first report on the biological control using by B. mojanvinensis to the lettuce Sclerotinia rot.

An antagonistic bacteria, Stenotrophomonas maitophilia BW-13 strain which was effectively inhibited mycerial growth of Bottom rot pathogen, Rhizoctonia solani PY-1 strain was isolated from the rhizosphere of the lettuce in Uiryeong-Gun, Gyeongsangnam-Do from 2002 to 2003. For the biological control, the most suitable inoculum and its density of pathogen, PY-1 strain ware tested prior biological control test, For the pathogenicity test, A inoculum (wheat bran)sawdust+rice bran+PDB) showing disease incidence of 100％ was selected as the most suitable inoculum, which showed more effective than B inoculum (sawdust＋rice bran＋DW) and mycelial disc. also, In selection of the amount of inoculum (40g, 50g, 60g, 70g, 80g), most suitable amount of inoculum of pathogen determined as 40g showing disease incidence of 80％. For the selection of effective microorganism to control bottom rot on lettuce, about 200 isolates were isolated from the diseased soil and lettuce leaves, and examined their antifungal activity to the pathogen on PDA. As the pots assay, BW-13 strain showed the highest control value as 90％, and followed by R-13 and R-26 strain as 80％ and 60％, respectively. Selected BW-13 isolates identified as 5. maltophilia (GeneBank accession no. AJ293473.1, 99％) by 16S rRNA sequencing. This is the first report on the biological control using by S. maltophilia to the bottom rot pathogen, Rhizoctonia solani PY-1 strain.

Ginseng (Panax ginseng) is one of the most widely cultivated medicinal herbs in Korea. During 3 or 5 years cultivation of ginseng, yield losses can reach as high as 30-60％ due to numerous diseases in Korea. Among 106 Bacillus strains isolated from various plant internal roots, we selected three promising biocontrol agents by screening against root rot caused by Cylindrocarpon destructan in a greenhouse. Preinoculation of selected isolates to seed or one-year-old root resulted in stimulation of shoot and/or root growth of seedlings, and control of root rot in infested soils with Cylindronrpon destructans (P=0.05). Furthermore, drenching of selected isolates on seedling-growing pots reduced the incidence of Phytophthora blight when the seedlings were challenged with zoospores of Phytophthora cactorum (P=0.05). However, isolates B1141 and B1142 did not show any antifungal activity against various soilborne pathogens while B1146 did in vitro. Our results provide an insight that rhizobacteria can induce resistance against various plant diseases on ginseng even if any resistant breeds have been unknown on ginseng yet.

To screen for potential biocontrol agents against postharvest disease of garlics caused by Penicillium hirsutum, a total of 933 isolates (432 fungi and 501 bacteria) were isolated from the rhizoshere or rhizoplane of garlics. Among them, Pantoea agglomerans isolate 59-4 (Pa 59-4) was selected for a potential biocontrol agent by in vivo wounded garlic bulb assay, When the spore suspension (10$\^$5/ spores/$m\ell$) of Penicillium hirsutum was co-inoculated with spore or cell suspension of each fungal or bacterial isolate on wounded garlics, the isolate highly suppressed disease development. Soaking garlic bulbs in the suspension of Pa 59-4 significantly reduced garlic decay from p. hirsutum. However, Pa 59-4 did not inhibit the mycelial growth of P. hirsutum in dual-culture with P. hirsutum on Tryptic soy agar. In order to elucidate mode of action of Pa 59-4 nutrient competition between Pa 59-4 and P. hirsutum was investigated using tissue culture plates with cylinder inserts containing defusing membrane reported by Janisiewicz et al. The results showed that Pa 59-4 effectively suppressed spore germination and mycelial growth of blue mold in the low concentration (0.5％) of garlic juice, but did not suppress those of blue mold in the higher concentration (5％) of garlic juice. This result suggests that the mechanism in biocontrol of garlic blue mold by Pa 59-4 may involve in nutrient competition with P. hirsutum on garlic bulbs.

Bacterial isolates, CC178, PTC25, HC39 and KY165 originally obtained from the leaves of cucumber or tomato were selected for biocontrol agents against gray mold of cucumber and tomato by in vivo cucumber seedling assay. Each suspension of the selected epiphytic bacteria were sprayed three times at seven-day interval from early stage of cucumber in a field. Incidence of gray mold on cucumber fruits treated with isolates CC178, PTC25, HC39 and KY165 was 15.3％, 18.2％, 23.6％, and 10.4％, respectively, whereas that of control was 38.0％ after 7 days of final spray. On the other hand, treatment with the selected isolates, CC178, PTC25, HC39, and KY165 on tomato showed 2.2％, 1.3％, 2.9％, and 3.5％ in the incidence of gray mold on leaves, whereas that of control was 9.3％. All selected isolates had strong antagonistic activity against Botrytis cinerea on dual culture plate assay.

The Plant growth Promoting rhizobacteria (PGPR), Pseudomonas sp. (A3) and Bacillus sp. (AT33) were isolated from the rhizosphere of Amaranthus blitum collected from soil contaminated with chromium. Both bacterial strains quantitatively reduced hexavalent chromium to trivalent chromium. Pseudomonas sp. broughter greater conversion of Cr6+ in the medium (100％) as compared to Bacillus sp.(62％). Phaseolus mungo seeds inoculated with Pseudomonas sp. or Bacillus sp. were grown under different concentration of chromium. The monitored parameters included elongation of shoot and root, fresh weight, dry weight and concentration of chromium in the shoot and root systems. As compared to non inoculated seedlings those inoculated with A3 and AT33 exhibited better growth.

The aim of the present study was to assess the importance of siderophore producing rhizobacteria on the growth of Brassica juncea under chromium stress. Pseudomonas sp. (A4) produced an iron chelating substance siderophores in iron deficient medium. Under chromium stress condition Pseudomenas sp. (A4) markedly increased the root and shoot length and also biomass of Brassica juncea as compared to Pseudomonas sp. (A3). This plant growth promotion has been related to the microbial production of siderophores.

Bacillus subtilis strain BAC02-4 was tested for its ability induced systemic resistance(ISR) in rice against Magnaporthe grisea We extend these studies to investigate the biological induction of systemic resistance in rice following treatment with the inducer isolate BAC02-4 and naturally infested with Pyricularia oryzae. We also determine levels of ISR activity during the period between disease development and the onset of systemic resistance. Comparition of lesion number according to applied concentration of BAC02-4 to 'Nagdongbyeo' when naturally infested with the conidia of P. grisea. Results from the blast nusery trial using the 'Nagdongbyeo' showed very low rice blast severity with the inducer concentration of 10$\^$8/ cfu level. Considering the low level of treatment and untreated control were observed to have developed typical susceptible lesion type. Highest protection against the rice blast pathogen when applied three times with 5 days interval as root drench at 5 to 6 leaf stage before pathogen challenge. But higher dose of bacterial inducer produced a little stunted plants with less number lesions and delayed disease development. Diseased leaf area of treated with suspension of the isolate which gave about 80％ of control efficacy at 20 days later comparable to that in noninfested, inducer-free soil.

The fermentation process and subsequent processing determine the efficacy of a bioherbicide propagule. Large batches of biomass of the mycoherbicide agent for white clover, Sclerotium sp.(BWC98-105) was produced in simple liquid fermentator in 5 gallons vessels(Model No. 8087, Dabo Inc., Korea) with oxygen supply(DPH16000, FineTech Inc., Korea) simulating industrial conditions by utilizing commercially available, inexpensive ingredients (10 ％ rice bran), The maximum biomass yield of Sclerotium sp.(BWC98-105) was obtained after 5 days of air pumped incubation at room temperature condition(22-28$^{\circ}C$). By using this simple facility, it could get fragmented or proliferated greatly and attained maximum mycelia biomass. The biomass of mycoherbicide agent consisted of hyphae devoid of spores. Biomass mycelia of the fungus 99％ survival at room temperature after 2 me. A thorough understanding of the effects of fermentation and formulation on viability and virulence is required to guide these processes. After an economical yield level of bioherbicide propagule has been achieved in a fermentation process, formulation becomes a critical factor which influences product efficacy. Because the fermentation must be stopped at a point when virulence/viability are optimum, the live bioherbicide propagule must be stabilized, formulated, and packaged.

Viral disease symptoms were investigated in the field grown Longiflorum hybrid cultivars, and the damages caused by infection with Lily mottle virus (LMoV) and Cucumber mosaic virus (CMV) were assessed by comparing growth of plants produced from seeds of Longiflorum hybrid cultivar both infected by artificial inoculation and free from infection with theses viruses. Dominant symptom caused by spotaneous infection with LMoV and CMV in the field was mottle combined with chlorotic stripe on leaves. LMoV developed brownish necrotic lesion on floral leaves. The incidence of viral disease by mixed infection with LMoV, CMV or Lily symptomless virus (LSV) in the filed grown Longiflorum hybrid cultivar, cultivated for more than 6 years, was 80 to 84 percent. In comparison with virus-free plants, plants doubly infected with CMV and LMoV by artificial inoculation decreased stem length by 14 percent and fresh weight by 38 percent. In conclusion, flower quality and the stem length of Longiflorum hybrid cultivar were affected by LMoV and CMV infection.

Soil metagenome is untapped total microbial genome including that of the majority of unculturable bacteria present in soil. We constructed soil metagenomic library in Escherichia coli using DNA directly extracted from two different soils, pine tree rhizosphere soil and forest topsoil. Metagenomic libraries constructed from pine tree rhizosphere soil and forest topsoil consisted of approximately 33,700 clones and 112,000 clones with average insert DNA size of 35-kb, respectively. Subsequently, we screened the libraries to select clones with antimicrobial activities against Saccharomyces cerevisiae and Agrobacterium tumefaciens using double agar layer method. So far, we have a clone active against S. cerevisiae and a clone active against A. tumefaciens from the forest topsoil library. In vitro mutagenesis and DNA sequence analysis of the antifungal clone revealed the genes involved in the biosynthesis of antimicrobial secondary metabolite. Metagenomic libraries constructed in this study would be subject to search for diverse genetic resources related with useful microbial products.

The Gyeonggi-do Agricultural Research and Extension Services has developed a web-site (www.epilove.com) in collaboration with EPINET to provide information on agricultural weather and rice disease and insect pest management in Gyeonggi-do. Weather information includes near real-time weather data monitored by automated weather stations (AWS) installed at rice paddy fields of 11 Agricultural Technology Centers (ATC) in Gyeonggi-do, and weekly weather forecast by Korea Meteorological Administration (KMA). Map images of hourly air temperature and rainfall are also generated at 309m x 309m resolution using hourly data obtained from AWS installed at 191 locations by KMA. Based on near real-time weather data from 11 ATC, hourly infection risks of rice blast, sheath blight, and bacterial grain rot for individual districts are estimated by disease forecasting models, BLAST, SHBLIGHT, and GRAINROT. Users can diagnose various diseases and insects of rice and find their information in detail by browsing thumbnail images of them. A database on agrochemicals is linked to the system for disease and insect diagnosis to help users search for appropriate agrochemicals to control diseases and insect pests.

From the PMMV (pepper mild mottle virus)-inducible ESTs differentially expressed in Capsicum chinense PI257284, we isolated a full-length cDNA (CcPHZF： Capsicum chinense phenazine), encoding a phenazine biosynthesis protein which catalyzes the hydroxylation of phenozine-1-carboxylic acid to 2-hydroxyphenazine-1-carboxylic acid. Phenazine compound has been known to exhibit broad-spectrum of antibiotic activity against various species of bacteria and fungus. The entire region of CcPHZF is 879 bp in length and the open reading frame predicted a polypeptide of 292 amino acids. The homolog of CcPHZF is not Present in database except clones of AC004044 and NM100203 from Arabidopsis with 58 and 59％, respectively. Genomic Southern analysis indicated that the pepper genome contains a single copy of CcPHZF. The CcPHZF was strongly induced in the pepper leaves 3 days after PMMV treatment, when HR occurs on the leaf surface. Characterization of CcPHZF is underway to investigate if the CcPHZF is related to disease resistance against pathogens.

A basic spray program for apple in which fungicides are scheduled to spray at 15-day interval from petal fall to late August was formulated on the properties of several selected fungicides. In order to improve it, experimental plots, completely randomized block with 3 replications, were prepared in an orchard of 15 years old Fuji cultivar, and the spray programs in which only one chemical in the basic spray program was substituted with others were applied to each plot. It was revealed that only single substitution of the fungicide in the basic spray program makes a great differences in the control of white rot and bitter rot, and that the control property of the fungicides against the two diseases was quite variable even by the time of application. A simila! ! r trial was conducted in 2002 with a new basic spray program that was formulated with fungicides that have shown best control in each spraying time in the previous trial, similar results were obtained. Applying this method, the usefulness of certain fungicide in the spray program for apple could be properly assessed. Anthracnose of Robinia pseudo-acacia L. caused by Collectotrichum spp.

Control effect of grafting with rootstocks on Phythophthora blight of pepper was evaluated. The 40∼45 day old seedlings of five pepper cultivars, Wangdaeback, Jinmi, Boosa, Samsungcho and Pochunngchun, as a scion were grafted with ten species of root stocks. There wasn't graft incompatibility in all scion-root stock combinations. After 21 days from inoculation with Phytopkhora capsici, incidence of Phythophthora blight of plant grafted with Kataguruma, R-Safe, R-16, YCM 334 and SCM 334 of rootstocks tested was decreased by 64.6∼100％ compared to non-grafted plant whose disease severity was 3.7. However, plant grafted with Umsung native and Yeongdong native pepper was more sensitive to Phythophthora blight than non-grafed plant. Control effect by grafting was inversely proportional to virulence of inoculum and not significantly different among scion cultivars used. And resistant reaction of scion against Phytophthora blight was not affect that of scion-root stock plant in this study.

The effect of biological control on the severity of hot pepper wilt disease was evaluated in the vinyl house with plants cultivated in the nursery soil containing chitin and chitinolytic microorganisms. The chitinolytic microorganisms, Trichoderma harzianum and Chromobacterium sp. strain C-61, were well survived in the nursery soil containing chitin. The hot pepper damping-off was markedly suppressed in the nursery soil containing chitin and chitinolytic microorganisms. The survival of chitinolytic microorganisms and suppression of damping-off were superior as the amounts of chitin added to the nursery soil increased, but growth of hot pepper was inhibited in the 10％ (w/w) chitin treatment. When the plants cultivated in the nursery soil containing 1％ chitin and chitinolytic microorganisms were transplanted in the vinyl house, the vegetative growth increased and the wilt disease was reduced as comparison with those of control.

The effect of biological control on the severity of hot pepper wilt disease was evaluated in the vinyl house with plants cultivated in the nursery soil containing chitin and chitinolytic microorganisms. The chitinolytic microorganisms, Trichoderma harzianum and Chromobacterium sp. strain C-61, were well survived in the nursery soil containing chitin. The hot pepper damping-off was markedly suppressed in the nursery soil containing chitin and chitinolytic microorganisms. The survival of chitinolytic microorganisms and suppression of damping-off were superior as the amounts of chitin added to the nursery soil increased, but growth of hot pepper was inhibited in the 10％ (w/w) chitin treatment. When the plants cultivated in the nursery soil containing 1％ chitin and chitinolytic microorganisms were transplanted in the vinyl house, the vegetative growth increased and the wilt disease was reduced as comparison with those of control.

Genetic diversity of Plasmodiophora brassicae from major chinese cabbage cultivating areas in Korea was analyzed by using PCR-RFLP and RAPD. Single spores of P brassicae isolated from galls of club root made induce lesion on chinese cabbage successfully. The PCR-RFLP and RAPD by primers PbITS, URP 3, 6 and OPA 7 revealed that single spore isolates showed various DNA polymorphisms among them unrelated geographic origins. These results indicate that P. brassicae population in Korea showed genetic difference among them. This study could be facilitate to identify genetic characteristics ofP. brassicae based on DNA polymorphisms between single spore isolates and to get basic information which can be used to advanced resistance breeding against club root of chinese cabbage.

Levels of blister rust infection (from Cronartium ribicola) varied in western white pine(Pinus monticola Dougl.) seedlings grown in two nurseries in northern Idaho. This observation suggested the potential importance of environmental components operating on the blister rust pathosystem. In an experiment designed to test the influence of environmental conditions at two nurseries, seedlings of a single genetic source were unintentionally held in cold storage for 6 months longer at one nursery than at the other. Subsequently, these seedlings, which had been growing under nursery conditions for 7 months or 1 month, were inoculated with blister rust spores on September 9th, 1999. Infection efficiency measured on the seedlings with only 1 month of growth was 70X greater than on the seedlings that had 7 months for their new growth to mature. Results from this nursery test and infection levels of northern Idaho resistant selections in mild climates suggest that expression of genes related to rust resistance in western white pine can be manipulated by regulation of phonology. If so, several new molecular tools may be employed to enhance our understanding of environmental regulation of genes for blister rust resistance.

More than 30 isolates of Alternaria were obtained from various solanaceous crops in Korea. For all isolates, morphological characteristics of the conidia were determined and compared with those of representative isolates of A. solani and A. tomatophila. Pathogenicity test was performed to Potato, tomato, egg plant and red Pepper and molecular characteristics of them including the representative isolates were determined using sequence analyses of ITS rDNA and histone H3 gene, and URP-PCR analysis. Based on morphological characteristics, the isolates from the solanaceous crops were grouped as identical or very similar to either A. tomatophila(ATO), A. solani(ASO), and unidentified Altemaria sp.(ASP). Among the molecular markers used in this study, the URP-PCR analysis was found to be appropriate for taxonomic resolution of these species. Based on the conidial morphology, pathogenicity test and molecular characteristics, A. tomatophila(early blight of tomato) could be distinguished from A. solani(early blight of potato), and the Alternaria sp.(ASP) from potato, which was closely related to A. solani in conidial morphology, was considered as a new species.

A potexvirus, Hosta virus X (HVX-Kr), causing mosaic and mottle symptoms was isolated from hosta plants (Hosta spp.), and its entire genome RNA sequence was determined. in Korea using cDNA library and RACE methods. The genome of HVX encodes five open reading frames coding for viral replicase, triple gene block (TGB), and viral coat protein (CP) from the 5'to 3' ends, which is a typical genome structure of potexviruses. The 3-terminal region of the virus includes the TGBI (26 kDa), TGB2 (13 kDa), TGB3 (8 kDa), and 23 kDa coat protein (CP) and the 3-nontranslated region (NTR). The CP gene of the type isolate of HVX (HVX-U) was amplified by RT-PCR and its nucleotide sequence was determined. The CPs of HVX-Kr and HVX-U had 100％ and 98.9％ identical amino acids and nucleotides, respectively. Most of the regions of the genome HVX had over 50％ nucleotide identical to other sequenced potexviruses. This is the first report of complete genome sequence information of HVX and molecular evidence supporting the virus as a distinct species of the genus Potexvirus.

Five pepper-infecting potyviruses, Pepper mottle virus (PepMoV), Chilli veinal mottle virus (CVMV), Pepper veinal mottle virus (PVMV), Pepper severe mosaic virus (PSMV) and Tobacco each virus (TEV), are known filamentous virus and can be infected pepper crops systemically. To understand pathology and genome information of the five viruses on pepper plants, host reactions and sequences were compared to the 5 viruses. Five potyviruses were inoculated onto some typical cultivars of hot peppers and compared their symptoms, and virus accumulations. A set of degenerate primers for potyviruses were applied to 5 viruses and RT-PCR was performed. RT-PCR products containing partial nuclear inclusion b and coat protein (CP) genes were cloned. Then, oligo dT primer and species-specific primer were redesigned to amplify the C-terminal part of CP and 3' noncoding regions of each viruses. Sequences of the viruses were analyzed and compared to serological relationships among the viruses. The data can be useful for screening of potyviruses in pepper plants and pathogen-derived transgenic pepper plant development.

A new isolate of rod-shaped virus was identified from grafted cactus, Gymnocalycium mihanovichii grafted onto Hylocereus trigonus, in Korea. The virus proved to be a new Tobamovirus and called previously as Tobamovirus-Ca for which we suggest the name Cactus mild mottle virus(CMMoV), because it produced systemic mild mosaic symptoms on its original host. CMMoV is distantly related to known species of the genus Tobamovirus on the basis of host range, serological and sequence analyses. Western blot analysis showed that CMMoV is serologically unrelated to Summons' Opuntia virus which is the only known species of the genus found in cactus plants. The 3'-terminal 2,910 nucleotides have been sequenced for the virus. The coat protein (CP) and movement protein (MP) genes encode 161 and 306 amino acids residues, respectively. The nucleotide and amino acid sequences of the CP were 39.6 ％ to 49.2 ％ and 26.4 ％ to 40.3 ％ identical to other tobamoviruses, respectively. The MP and 3' noncoding region shared 16.3 ％ to 23.3 ％ and 44.6 ％ to 63.4 ％ identities, respectively, with the members of the genus. Phylogenetic tree analysis of the CP gene revealed that CMMoV clusters with members of subgroup I of Tobamovirus. CMMoV particles contained genomic RNA along with two subgenomic RNAs, and this characteristics is common in the members of the subgroup II. This is the first information of sequence and comparative analysis of a Tobamovirus that infects cactus.

Rice stripe virus causes severe damage to rice in Korea, Japan and China. RSV is a type member of the tenuivirus group and transmitted by the small brown planthopper, Laodeiphax striatellus, in a persistant manner. Until now, occurrence of RSV is limited in of southern part of Korea. But recently occurrence of RSV is increasing and spreading in central part of Korea including Chungcheong and Kyonggi province. So we analyzed recent occurrence trend of RSV which is increased and cloned and sequenced coat protein gene for isolating of RSV strain. Infected rice of each species(Ilpumbyeo, Sindongjinbyeo, Keumobyeo-2, Dongjinbyeo, Jongnambyeo, Samcheonbyeo, etc.)is collected, we extracted total RNA from infected leaves and detected RSV viral RNA by reverse transcription(RT)-PCR using specific primer of coat protein gene. The result of RT-PCR, we observed specific band. We already cloned cDNA from the band, is analyzing sequence variety and homology of each species.

Post-harvest decay, caused by Penicillium spp. is a serious problem of fruits worldwide. Morphological characteristics and molecular markers were used to characterize 22 Penicillium isolates from apples, 18 isolates from pears, 60 from oranges and 18 from grapes and 23reference isolates representing related Penicillium spp. to assess their diversity and resolve their taxonomy. Based on morphological and physiological characteristics, the isolates were grouped as identical or very similar to P. digitatum, P. italicum, P. ulaiense or very similar to P. crustosum, P. expansum, P. solitum and unidentified Penicillium spp. Based on sequence comparisons of ITS region, variable site were presented within and among the species, but there variation were not correlated with the species. Cluster analyses of AP-PCR fragment patterns using UP and L45 primer and the -tubulin gene sequence, the Penicillium species were segregated into distinct groups. Particularly. the -tubulin partial sequence data provided support for species concepts based on morphological and physiological characteristics.

Complete genomic nucleotide sequence of Daphe virus S (DVS), a member of the genus Carlavirus, causing leaf distortion and chlorotic spot disease symptoms in daphne plants, has been determined in this study. The genome of DVS contained six open reading fames coding for long viral replicase, triple gene block, 36 kDa viral coat protein (CP) and 12 kDa from the 5' to 3' ends, which is a typical genome structure of carlaviruses. Two Korean isolates of DVS isolates were 98.1％ and 93.6％ amino acid identical in the CP and 12kDa, respectively. The CP gene of DVS shares 25.2-55.2％ and 42.9-56.1％ similarities with that of 19 other carlaviruses at the amino acid and nucleotide levels, respectively. The 3'-proximal 12 kDa gene of DVS shares 20.2-57.8％ amino acid identities with that of 18 other members of the genus. The 3' noncoding region of DVS consists of 73 nucleotides with long excluding poly A tract, and shares 69.1-77.1％ identities to the known carlaviruses. In the phylogenetic analyses of the two proteins, DVS was closely related to Helenium virus S and Chrysanthemum virus B. This is the first complete sequence information for the DVS, and further confirms the classification of DVS as a distinct species of the genus Carlavirus.

Fusarium oxysporum f. sp. fragariae is a fungal pathogen causing wilt disease on strawberry. The RAPD and RFLP of IGS region of rDNA were used to identify genetic affinity among 22 isolates of F. oxysporum f. sp. fagariae obtained from various location of major strawberry cultivating areas in Korea. Approximately 2.6kb DNA fragment was amplified with primer CNS1 and CNL12, and polymorphisms were observed with Avail and HinfI. A dendrogram was constructed using the UPGMA for cluster analysis. Eight distinct groups were clustered based on the banding pattern obtained from RAPD and rDNA RFLP. There was high level genetic variation among Korean isolates of Fusarium of sporum f. sp. fragariae.

Late blight, caused by Phytophthora infestans, is one of the most destructive disease on potato and tomato cultivation. To analysis genetic diversity P. infeatans isolates were collected from potato and tomato fields in Korea. These pathogens contained both Al and A2 mating type with metalaxyl-resistant and sensitive isolates. Polymorphisms showed base on RAPD (Random Amplified Polymorphic DNA) in both potato and tomato isolates of P. infestans. Cluster analysis showed high level genetic variation in potato isolates of P. infestans than tomato isolates. P. infestans isolates were observed genetic diversity among them but not grouped among isolates related mating type and metalaxyl response. These results exhibited that P. infestans isolates showing genetic difference among them were distributed in Korea.

Phytophthora blight, caused by P. capsici, is very important disease of pepper. Many fungicides to control of Phytophthora blight have been developed, but most of fungicides disappeared in short periods. Nowadays dimethomorph was known as one of the most effective to control of this disease. P. capsici isolates from pepper fields were collected and surveyed their growth in dimethomorph amended V8 medium in order to evaluate their fungicides resistance. The fungicide resistant isolates were not founded among them. Most of the sensitive isolates were inhibited perfectly in V8 medium amended with 10ppm dimethomorph. Mutants of P. capsici by dimethomorph, was grown very well in 250ppm. The difference of pathogenicity, colony morphology, drug response, RT-PCR results was identified between sensitive and resistance isolates. This study should be provided a basic information about the occurrence of dimethomorph resistant isolates and genetic changes in P. capsici population.

The importance and diversity of the genus Stemphylium highlights the need for accurate identification of species. However, many Stemphylium isolates have been misidentified due to the use of spore size as the only identifying character. Molecular phylogenetic analyses were performed on fifty-four isolates covering 9 Stemphylium species collected in Korea. Phylogenetic analysis of the translation elongation factor -1 alpha (EF-1) and the calmodulin gene sequence data showed that Stemphylium species were segregated into seven distinct groups, most of w hichcorrelated with species identified by morphology. Analysis of EF-1 in particular was useful for establishing well- supported relationships among the species of Stemphylium.

We tested for the presence of double-stranded RNA (dsRNA) mycovirus in 827 Fusarium graminearum isolated from diseased barley and maize. dsRNA mycoviruses with various sizes were isolated. Of them, it was previously reported that dsRNA from DK2l isolate had pronounced morphological changes, including reduction in mycelial growth, increased to red pigmentation, reduced virulence and sporulation. (Chu et al., Appl. Environ. Microbiol. 2002). For better understanding of this hypovirulence associated with DK2l dsRNA virus, we determined the complete nucleotide sequence of dsRNA genome and named Fusarium hypovirus DK2l strain (Fhv-DK2l ). Genomic RNA of Fhv-DK2l was determined to be 6625 nucleotides in length excluding the poly (A) tail and contained three putative open reading frame. RNA-dependent RNA polymerase (RdRp) and helicase domain were expected in ORF A, 54 to 4709 nucleotide position. ORE B, 4752 to 5216 nucleotide position, and ORF C, 5475 to 6578 nucleotide position, were predicted to encode 16.7kDa and 41.3kDa protein respectively each. We could not detect any conserved domains from these two proteins. Phylogenetic analysis showed Fhv-DK2l was related to Cryphonectria hypovirus 3. Ten additional isolates were found that were infected with dsRNA mycoviruses. These mycoviruses contain 2 to 4 different segments of dsRNAs with the size range of approximately 1.7 to 10-kbp in length. The presence of dsRNAs isolates did not affect colony morphology and were transmissible through conidia and ascospore with incidence of 30-100％. These results indicate that there is genomic diversity of dsRNA mycoviruses that infect F. graminearum isolates and that impact of virus infection on host's morphology and virulence is determined by the interaction between dsRNAs and the fungal host, not by the mere presence of the dsRNAs

The glyoxysomal nature of microbodies was determined in Botryosphaeria dothidea hyphae based on morphology and in situ enzyme characteristics by transmission electron microscopy and cytochemistry. Bound by a single membrane, microbodies had a homogeneous matrix and varied in size ranging from 200 to 400 m in diameter. Microbodies had crystalline inclusion(s) which consisted of parallel arrays of fine tubules in their matrices. Microbodies and lipid globules were frequently placed in close association with each other, forming microbody-lipid globule complexes in hyphae. The cytochemical activities of catalase and malate synthase were localized in matrices of microbodies, showing intense electron-density of the organelle. In addition, the immunogold labeling detected the presence of catalase in multivesicular bodies and hyphal cell walls as well as in matrices and crystalline inclusions of microbodies, supporting the enzyme secretion through cell walls. Meanwhile, isocitrate Iyase was localized only in matrices of microbodies. These results suggest that microbodies, particularly complexed with lipid globules, in the fungal hyphae are functionally defined as glyoxysomes, where glyoxysomal enzymes are biochemically active for the glyoxylate cycle to be a metabolic pathway in gluconeogenesis. (Mycology and Fugus Diseases)

Seed-borne fungal pathogens on rice seeds were investigated in order to evaluate their effect on rice grain quality. Rice seeds of two cultivars, Ilmibyeo and Daesanbyeo were collected from 27 areas of Korea and the fungal parasites on seeds were isolated by using a blotter method. Isolation frequency and number of species were varied from collection areas of seed samples. A total 13 species of fungi were identified from the seeds. Among them, Altemana alternata was the most frequent over the country. Bipoiaris oryzae most commonly from Gyeongbuk and Jeonbuk, Alternaria padwickii from Jeonbuk, and Nigrospora oryzae from Choongnam. However Bipolaris. oryzae, Alternaria padwickii, and Nigrospora oryzae were the most frequently isolated fungi from Gyeongbuk and Jeonbuk, and Chungnam, respectively. B. oryzae, A. alternata, A. padwickii, and N. oryzae were dominants on Ilmibyeo showing 10.3％, 10.2％, 5.2％, and 5.2％ infection rate, respectively. While, N. oryzae, A alternata, and Cladosporium sp. were most frequently isolated fungi from Deasanbyeo revealing 15.1％, 9.6％, and 7.5％ infection rate, respectively. These fungi inhabiting on hulls or endosperms of rice seed might be considered as potential factors decreasing rice grain quality. Further investigation of the fungi on grain rice quality are undergoing.

White rot on Allium species recently had a high incidence as increased cultivating areas of tropical garlic types in Korea. Two types of Sclerotium have been known as causal agents producing different size and shapes of sclerotia in infested fields. We developed a new method for isolation of two types of sclerotia from infested field soils that can be used for ecological study of sclerotium spp. and establishment of control strategy. Soil samples collected from heavily infested fields were evenly mixed and placed on a automatic sieve shaker connected with tap water. After 10 min. of shaking, residues on 0.5mm and 0.25mm sieve were separately collected and suspended with 70％ sugar solution, which method floats sclerotia in aqueous layer. Then, floated fraction was carefully separated and mixed with a same volume of 1％ sodium hypochlorite solution to differentiate with organic materials. This method provides direct count of sclerotia under dissecting microscopy.

Agrobacterium vitis is a causal agent of crown-gall disease on grapevine. In Korea, grapevine variety (GeoBong) have severely been infected by the bacteria since stems of the variety were buried in soil for overwintering. Infection ratio over 70-80％ was observed on 7 years old GeoBong grapevine in Ansung and Cheonan. PCR specific primers for A. vitis strains were designed using nucleotide sequences of vir A gene in Ti-Plasmid, pheA gene in chromosomal DNA and a URP-PCR polymorphic band. Three hundred bacterial strains were isolated from the different 80 galls formed on GeoBong grapevine in Cheonan and Ansung of Korea and were screened to identify A. vitis using the three specific PCR primers for Agrobacterium vitis. Twenty-four bacterial strains that are detected by the primers were further confirmed by pathogenicity and biochemical methods. To investigate the genomic diversity of the bacterial strains, twenty primers of 20 mer referred to universal rice primers (URP) were applied for PCR fingerprinting, Of them, URP2R and URP2F primers could effectively be used to detect polymorphism within the bacterial strains.

Relative degree of resistance of citrus to Xanthomonas axonopodis pv. citri, the causal bacterium of canker, was investigated. Growth rate of a bacterium in leaf tissues after infiltration, disease incidence, and percent of lesion area were compared. By using growth rate[(GR=(At - A$\sub$t-1/)/A$\sub$t-1] host plants were differentiated into susceptible and resistant. Growth rates reached to peak at 40 hrs after inoculation and then declined. The growth rate in leaf tissues of a moderately susceptible cultivar, Citrus sinensis vu. Lane late(sweet orange), was the highest, and those of C. unshiu ${\times}$ C. sinensis(kiyomi), C. junos(yuzu), [(Citrus. unshiu x C. sinensis) x C. reticulata] (shiranuhi), and C. unshiu(satuma mandarin) were similar. This result indicates that the growth rate of the bacterium in leaf tissues can be effectively used for evaluation of disease resistance for citrus plants to X. axonopodis pv. citri. The disease on sweet orange occurred earlier than relatively resistant citrus plants tested. The percent of lesion area on leaf was also higher in sweet orange than those of satsuma mandarin, shiranuhi and kiyomi, and yuzu. The disease severity was highest on sweet orange and followed by kiyomi, shiranuhi, satsuma mandarin, and yuzu.

Dispersal of Xanthomonas axonopodis pv. citri, causing citrus bacterial canker disease on Unshiu orange was investigated at previously infested plots at Seogwipo in Jeju island of Korea. The bacterial pathogen overwintered in lesions started to multiply at tate May, and disease firstly observed one month after detection of phage from lesions. The disease gradually increased, however, it dispersed non-directionally to nearby plants from inoculum sources. Diseased plants were aggregated to form a cluster throughout the experiment. Population dynamics of phage on symtomless leaf surface and the disease severity were compared in the nursery, Increase of phage population on symptomless leaf surface preceded one month to that of the disease severity Population of phage increased constantly from late July to October, however, the disease severity decreased from late August to late October. It was assumed that the decrease of disease severity might be due to disease-induced defoliation.

Since the demonstration that the transgenic plants expressing tobacco mosaic virus(TMV) coat protein(CP) gene showed resistance to TMV infection, there have been numerous attempts to produce virus-resistant plant by introducing of a part of or modified viral genome. This study was conducted to investigate the characterization and variability of disease outbreak of transgenic potato(T-potato) with the CP gene of potato leaf roll virus(PLRV) in an isolated field from 2000 to 2002. In the field inspection, incidence of PLRV on T-potato showed only 3.5％, while non-transgenic potato(N-potato) revealed 13.4％. Infection rate of PLRV was considerably low on T-potato with 4.2％ compared to 15.4％ of N-potato in ELISA tests. Those of potato virus M, potato virus Y and potato virus X on both potatoes were not statistically different. Infection of potato virus A was not observed on both potatoes. Incidence of potato late blight caused by Phytopkhora infestans on T-potato and N-potato did not differ each other with 52.7％, and 50.8％, respectively, Mating type of the causal fungus isolated from both potatoes was all Al types. Results indicates that the CP gene of PLRV affects specifically to the virus in the transgenic potato.

The objective of this work was to analyze the population of sequence variants of citrus tristeza virus (CTV) isolates in Korea and to make the phylogeny trees of CTV in Korea. We also tried to analyze and find the mild strain of CTV to apply for the cross protection. The CTV isolates from yuzu (C. Junos) collected from different geographic areas of Southern provinces such as Namhae-Do, Kerche-Do, Bosung, Wan-Do and Koheung and Jeju-Bo, Korea were used for SSCP analysis. The SSCP profiles of the cDNAS obtained by RT-PCR with primers specifically designed for the p20 of the CTV population. The SSCP profiles obtained from 150 PCR products in yuzu contained two or three DNA bands, whereas, in some case, others contained four or more bands of similar intensity. The pathologically mild isolates of CTV usually yielded two DNA bands by SSCP profiles, whereas the SSCP profiles of the most virulent isolates contained more than two DNA bands. Plants shown severe stem pitting were corresponded to those plants with typical SSCP profiles of severe strains, and vice versa. This results indicate that the primers designed for SSCP analysis can be used for distinguishing the mild strains from severe strains of CTV.

Fusarium graminearum (telomorph：Gibberella zeae), an important fungal pathogen of cereal crops with ubiquitous geographic distribution, produces mycotoxins on diseased crops that has threaten human and animal health. Recently severe epidemics of scab diseases of barley and rice by this fungus occurred in Korea, causing serious economic losses. To determine genetic diversity of F. graminearum from rice in Korea, a total of 269 isolates were obtained from Southern part of Korea during 2001-2002. A phylogenetic tree of the isolates was constructed by using amplified fragment length polymorphism (AFLP). Population structure of the rice isolates consists of a single lineage (lineage 6). Frequency of female fertility among these Isolates was relatively low (37％) compared to that among lineage 7 isolates from Korean corn. PCR amplification using chemotype specific primers derived from Tri7 and Tri13 genes at the trichothecene biosynthesis gene cluster revealed that most isolates (260) were NIV chemotype；9 isolates were identified as DON chemotype by Tri13 but as either NIV chemotype or unknown by Tri7. The result of chemical analysis also supported the chemotype determination；all of the NIV chemotype isolates produced NIV, whereas the 9 isolates produce either DON or no toxin.

Pepper anthracnose is one of the major limiting factors in pepper production. Boring last over 10 years, Colletotrichum gloeosporioides has been known as the most prevalent species among five Colletotrichum spp. involved as anthracnose causing agents. Recently, however, the change of major species with pepper anthracnose has been proposed. Identification study was peformed on 12 test isolates collected from anthracnose disease symptoms on pepper during 2001-2002 and 25 reference isolates obtained from several other host plants. The identification of the isolates with morphological observation and IfS region sequence comparison resulted that 11 ones from 12 test isolates colleted from pepper anthracnose during 2001-2002 were identified as C. acutatum. PCR using species-specific primers designed from ITS region sequence suggested a rapid diagnosis method in identifying C. acutatum from C. gloeosporioides.

Since 2000, severe rots on aerial and underground parts of paprika( Capsicum annum L.) has occurred in most cultivation glasshouses throughout the country. Totally 169 isolates of a fungus were consistently isolated from the diseased plant tissues of fruits, stems, branches, and roots collected from 19 farms in six provinces. Anamorph stage of the fungus was identified as Fusarium solani based on morphological characteristics. However, the fungus readily produced sexual structure of perithecia on infected plant tissues and on agar medium. Since the fungus formed abundant perithecia by single isolate, it was considered as a homothallic strain of Nectria hematococn, the teleomorph of F. solani. Irregularly globose perithecia with orange to red color formed sparsely to gregariously on dead tissues of fruits and basal stems at the 3ate infection stage, which is a diagnostic sign for the disease. Abundancy of perithecium varied among isolates and they sized 125-220$\mu\textrm{m}$ in diam. Asci enveloping eight ascospores were cylindrical and measured 60-80x8-12$\mu\textrm{m}$. Ellipsoid to obovate ascospores are two-celled and measured 11-l8${\times}$4-7$\mu\textrm{m}$. Ascospores are hyaline, slightly constricted at the central septum, and revealed longitudinal striations that is a typical trait of the species. This fungus that has never been reported in Korea previously became a threat to paprika cultivation because of its strong pathogenicity and nationwide distribution.

A near full-length sequence of a new tobamovirus infecting Hibiscus rosa-sinensis L. was determined. The genome consists of 58 nucleotides (nt) 5' UTR, followed by a 4.9 kb ORF which methyl transferase helicase domain (128 kDa), readthrough protein RNA dependent RNA polymerase (RdRp) 185 kDa and a 52 kDa protein. The 128 kDa protein had a maximum homology of 51.4 ％ to TMGMV and amino acids (an) were 54.3 ％ identical to TMV- vulgare strain. The 185 kDa RdRp had a maximum homology of 53.5％ to TMV-Ob and KGMMV-Y and a 59.6％ homology at the an level to CGMMV-SH. The MP gene encodes 282 aa and its theoretical molecular weight is 30.4 kDa. The nt and an sequence identities of MP ranged from 38.8％ to 43.9％ and 30.9％ to 37.9％, respectively. The CP gene encodes 163 residues and with a theoretical molecular weight of 18.2 kDa The (nt) and aa sequences of the CP were 46.9 ％ to 51.6％ and 45.3％ to 57.1％ identical to other tobamoviruses, respectively. The predicted virion origin of assembly (OAS) was located in the CP gene. Phylogenetic trees generated based on the nt and as sequences of RdRp, MP and CP genes indicated that this new virus clustered with subgroup II tobamoviruses. Although the CP ORF of this virus shared a high nt and aa sequence identity with Sunn-hemp mosaic virus (SHMV), Western analysis showed that it is serologically unrelated to SHMV. We propose the name Hibiscus virus S (HVS) for this Singapore isolate. This is the first report on a near full-length sequence of a Tobamovirus that infects hibiscus.

We describe two new Trichoderma species associated with oyster mushroom in Korea. Trichoderma green mould has been one of the most serious diseases of oyster mushroom in Korea. Of these the predominant species are two unrecorded species. We designed as Trichoderma sp. Korean type 1 (Th K1) and Trichoderma sp. Korean type 2 (Th K2), respectively. Th K1 and Th K2 can be distinguished from previously reported Trichoderma species as well as each other in morphological characteristics including growth rate at 35$^{\circ}C$, colony morphology, conidia shape and branch pattern of phialides. Sequence of the ITS region of rDNA, the protein coding translation elongation factor gene(EF-1${\alpha}$), and RNA polymeraseII (RPB2) not only clearly separated Trichoderma sp. Korean types from their closely related T. harzianum biotype but also distinguished them from each other. Analyses of the EF-1${\alpha}$ and RPB2 sequences were found to be more useful for establishing systematic relationships among Trichoderma isolates than those of the ITS sequence. Based on the results of morphological and molecular characteristics. We propose the two Trichoderma sp. Korean types as the new species

YMV-K strains were purified from D. oppostita Thunb. tv. Dung-Gun-Ma showing mosaic symptom on their leaves. YMV-K strains were filamentous particles of 780nm in length and induced cytoplasmic disorder such as inclusion body formation. Nucleotide and amino acid sequences of 5'-UTR, Pl and CP of YMV-K strains shared 80.8, 64.7 and 98.3％ identity respectively to JYMV J1 in the mean value. Purification of YMV-K strains according to JYMV purification method(S. Fuji) was conducted to product antiserum. With antiserum against YMV-K strains, the Various diagnosis methods such as IC-RT-PCR, DIBA, RIPA and indirect-ELISA were used to detect YMV-K strains in Chinese yam plant.

A leaf blight disease was found on peony plants growing in the first author's apartment garden in May, 2003. A species of Phytophthora was isolated from the lesions. The isolate readily produced sporangia and sex organs on V8 juice agar plates. Sporangia were papillate, ovoid to subspherical and caduceus with a pedicel. Sporangia were 33.6-38.4 ${\times}$ 33.6$\mu\textrm{m}$ with 1/b ratio approximately 1.14, papillae 4-5$\mu\textrm{m}$ high, pedicels also 4-5$\mu\textrm{m}$ long. Oogonia were spherical, 28.8$\mu\textrm{m}$ in diameter. Antheridia were globose, 14.4$\mu\textrm{m}$ in diameter and mating with oogonia paragynously. Mycelia grew best at 30$^{\circ}C$ and did not grow at 35$^{\circ}C$ or above, and at 5$^{\circ}C$. The morphological characteristics conformed to P. cactorum.

A leaf blight disease was found on Fatsia japonica plants growing in the first author's apartment garden in May, 2003. Major symptoms were leaf blight and petiole rot. A species of Phytophthora was isolated from the lesions. The isolate readily produced sporangia and sex organs on V8 juice agar plates. Sporangia were papillate, ovoid to subspherical and caducous with a pedicel. Sporangia were 33.6-38.4 ${\times}$ 33.6$\mu\textrm{m}$ with 1/b ratio approximately 1.14, papillae 4-5$\mu\textrm{m}$ high, pedicels also 4-5$\mu\textrm{m}$ long. Oogonia were spherical, 28.8$\mu\textrm{m}$ in diameter. Antheridia were globose, 14.4$\mu\textrm{m}$ in diameter and mating with oogonia paragynously. Mycelia grew best at 30$^{\circ}C$ and did not grow at 35$^{\circ}C$ or above, and at 5$^{\circ}C$. The morphological characteristics conformed to P. cactorum (Leb. And Cohn) Schroeter.

Shoot blight was occurring on shoots of suckers of common lilac (Syringa vulgaris L.) growing in first author's apartment garden in May 2003. A species of Phytophthora was isolated from the lesions. The isolate did not sporulate on agar media but formed sporangia in water and also formed sex organs in single culture. Sporangia were semipapillate, ovoid obpyriform, measured 45.6-52.8 ${\times}$ 33.6-36.0$\mu\textrm{m}$. Sporagia were very variable in shape. Optimum temperature for mycelial growth was 25$^{\circ}C$. Oogonia were spherical and antheridia were paragynous. Optimum temp for mycelial growth was 25$^{\circ}C$. The isolate was identified as Phytopkhora citricola on the basis of the morphological characteristics and cardinal temperature.

A leaf blight disease caused by a species of Phytophthora has been observed on castor bean plants growing near dwelling houses in Manchon-dong, Daegu since 1993. The first isolate that we have kept was producing papillate, ovoid-obpyriform to obpyriform sporangia with on a simple sympodial sporangiophore from diseased tissue placed on water agar plates. The pure isolate, however, did not sporulate on agar media, and rarely even in water, but produced mycelial swellings and chlamydospores in water. Sporangia measured 26.1-77.4 ${\times}$ 23.2-44.0$\mu\textrm{m}$. Chlamydospores were either terminal or intercalary, and measured 24-29.4$\mu\textrm{m}$ in diameter. Sex organs were not formed in a single culture. In 2003, another pure isolate was isolated from castor bean plants with similar symptoms at the same place. The second isolate was distinct from the first one in that the second isolate was readily and abundantly sporulating on V8 juice agar plates. Sporangia of the second isolate were papillate, ovoid and caduceus with a pedicel. Sporangia measured 19.5-48.8 x 17.6-34.3$\mu\textrm{m}$ with 3.7$\mu\textrm{m}$ high papilla and 4.1$\mu\textrm{m}$ long pedicel. No sex organs were formed in a single isolate culture. Both isolates were pathogenic on castor bean plants. Results of the efforts to identify the two species of Phytophthora will be discussed.

A destructive stem rot of strawberry (cv. Akihime) occurred sporadically in farmers' fields around Daegok-Myeon, Jinju City, Gyeongnam province in Korea. The infected plants showed stem and crown rot, sometimes whole plant blighted. White mycelia spread over stems of infected clones and sclerotia formed on the old lesions near to soil surface. The fungus formed white colony on PDA and showed maximum mycelial growth and scleotial formation around 30$^{\circ}C$. The fungus usually have many narrow mycelial strands in the aerial mycelium and the width were 4.0∼10.0$\mu\textrm{m}$. The typical clamp connections were formed on the mycelium. The shape of sclerotia was globoid and 1.0∼2.8 mm in size. The fungus was isolated repeatedly from the infected tissues and identified as Sclerotium rolfsii. The fungus was inoculated to strawberry and confirmed its pathogenecity This is the first report on the stem rot of strawberry caused by Scierotium rolfsii in Korea.

Tomato soil pathogens(Phythopthora spp.) analyed high rates in series culture soil and existed in culture parts. To make a diagnosis of Phythopthora sp. and Its concentration, potato slices were manufactured to a round shape(2.5cm) or retangular form(1x4cm). and then, The potato slices dipped into diagnostic reagents with an antibiotic substance for 2∼4hours. Potato slices treated with a few reagents varied into 15cm depths in innoculated soils for 24hrs. Mycelium of the Phytophthora root rot fungus, Phythopthora capsici, were produced easily on potato slice. We collected many potato slice samples on diseased fields in various area. After storage of 24hrs in 20$^{\circ}C$ incubator, White mycelium of Phythopthora sp. formed on potato slice surface. Dilute concentrations of Phythopthora sp. was detected very low contents(1${\times}$10$^1$sporangia/g). But expressing Phythopthora root rots on potato slice did not developed larger lesions upon storage time in room temperature. These results suggest that the use of potato slice in a series of soil cultural system may still serve as efficient means of diagnosis of Phythopthora root rots in the absence of control measures.

This study reports the identification of species of Colletotrichum strains originating from red-pepper and Chinese matrimony vine in Cheongyang. Ninteen isolates of red-pepper and 26 Coiletotrichum isolates of Chinese matrimony vine were compared with 5 isolates of strawberry representing C. gloeosporioides, by use of morphological and cultural criteria. Twenty three isolates among 26 isolates from Chinese matrimony vine were identified as C. acutatum, characterized by the low growth rates and the low sensitivity to carbendazim and diethofencarb. Also, all the isolates of red-pepper were identified as C. acutatum, showing the same characteristics as those of Chinese matrimony vine. Three and five isolates from Chinese matrimony vine and strawberry, respectively, were identified as C. gloeosporioides, characterized by the high growth rates and the high seneitivity to carbendazim and diethofencarb. There were differences in colony color and pathogenicity between Chinese matrimony vine isolates and red-pepper isolates of C. autatum. The isolates of C. acutatum from Chinese matrimony vine producing orange colored colonies with abundant spores showed the strong pathogenicity to Chinese matrimony vine, although they could not infect fruits of red-pepper by the wound inoculation. However, red-pepper isolates of C. acutatum producing gray colonies showed the strong pathogenicity to Chinese matrimony vine as well as red-pepper. Furthermore, comparative study on PCR amplification of ITS regions of rDNA was carried out using a number of Colletotrichum isolates. A species-specific primer could be used for the identification of C. acutatum from red-pepper and Chinese matrimony vine.

White rot on garlic caused by Sclerotium cepivorum firstly occurred at Goheoung, Jeonnam in 1998. Thereafter, the disease rapidly spread throughout the country except Gangwon and became a major limiting factor for the cultivation of various Allium species such as garlic, onion, and welsh onion. The disease that has not been reported on a wild garlic(Allium monanthum) previously occurred severely at Seosan, Choongnam in 2003. Among cultivation areas in the region, 10.7％ were infected by the disease and the ratio of diseased plant reached up to 55.0％ in some heavily infected fields. Two species of Sclerotium were consistently isolated from infected samples and identified as S. cepivorum or another Sclerotium sp. Averaged size of sclerotium of the former was 455.0x562.2 urn, while the later was 374.4${\times}$347.2$\mu\textrm{m}$. Patogenicity to Allium species and mycological characteristics such as sclerotium size, growth temperature, and microconidia of the fungi were similar to those reported on other Allium species previously. Consequently, the wild garlic is a newly reported host of the two pathogenic fungi in Korea.

A Phoma sp. was detected on rice seeds and was identified as Phoma sorghina (Sacc.) Boerema, Dorenbosch ＆ Van Kesteren based on their morphological and cultural characteristics. On oatmeal agar, pycnidia were abundant, solitary, scattered or gregarious, subglobose to flask-shaped, usually with a distinct neck and ostiole, glabrous, blackish-brown, 55∼185 ${\times}$ 40∼170 $\mu\textrm{m}$ in size. Conidiogenous cells were monophialidic, hyaline, subglobose to ampulliform, 3∼5 $\mu\textrm{m}$ in diameter. Conidia were ovoid to ellipsoidal, hyaline, unicellular and measured 3.5∼6.0 ${\times}$ 1.5 ∼3.0 $\mu\textrm{m}$ (usually 4.0∼5.0 ${\times}$ 2.0∼2.5 $\mu\textrm{m}$) in size. Chylarnydospores were variable, uni- or multicellular, intercalary or terminal, solitary or in chains, dictyosporous or botryoid. NaOH spot test was positive on malt extract agar. This is the first report of P. sorghina identified on rice seeds in Korea.

Anthracnose occurred on rapsberry grown in Gochang areas of Korea in 2003. The disease incidence was ranged from 1.1 to 2.6％. Anthracnose of rapsberry appeared as dark brown circular spots on naturally infected stems. The symptoms of infected stems were small brown to dark brown spots and gradually enlarged larger cylindrical dark brown lesions. The causal fungus of anthracnose isolated from the diseased plants was identifed as Colletotrichum coccodes based on the morphological and cultural characteristics. All isolates of C. coccodes were produced similar symptoms on the host leaves by artificial inocultion.

Leaf blight of kudzu ( Pueraria lobata ) was found in Jeonbuk province in 2002. The main symptoms appeared as leaf blight and showed yellowing and wilting. The causal pathogen of the leaf blight was isolated from symptomed kudzu leaf and produced white to cream, usually floccose mycelium. It readily formed reddish orange mycelium on PDA. It produced typical microconidia and macroconidia. The microconidia were the reniform. The macroconidia were wide, slightly curved, usually 3 to 4 septate and size was 45 ∼ 85 ${\times}$ 5 ∼ l0$\mu\textrm{m}$. The pathogen produced chlamydospore singly on short hyphal branches within 2 to 3 weeks, which was hyaline, globose, and smooth walled. The pathogen was, therefore, identified as Fusarium solani based on cultural and morphological characters. This is the first report on the leaf blight of kudzu caused by Fusarium solani in Korea.

Gummy stem blight, caused by Didymella bryoniae occurs exclusively on cucurbits. This fungus has been known not to produce its pycnidium in vitro unless irradiated. Through this study, we optimized cultural conditions for mass-production of pycnidiospore by Metal Halide Lamp irradiation. In brief, the mycelial was cultured at $26^{\circ}C$ on PDA, for 2 days under the darkness, and then the plate was illuminated with MH lamp continuously for 3-4 days at $26^{\circ}C$, a great number of pycnidia was simultaneously formed. Thus produced pycnidiospores were used as immunogen. From fusions of myeloma cell (v-653) with splenocytes from immunifed mice were car ried out. And, two hybridoma cell lines that recognized the immunogen Didymella bryoniae were obtained. One Monoclonal Antibody, Db1, recognized the supernatant and the other monoclonal antibody, Db15, recognized the spore. Two clones were selected which were used to produce ascite fluid two MAb Db1 and Db15, were immunotyped and identified as IgG1 and IgG2b, respectively. Titer of MAb Db1 and MAb Db15 was measured absorbance exceeded 0.5 even at a $10^{-5}$ dilution. The MAbs reacted positively with Didymella bryoniae but none reacted with other of fungi and CMV, CGMMV Sensitivity of MAb was precise enough to detect spore concentration as low as $10^{3}$ well by indirect ELISA characterization of the MAb Db1, Db15 antigen by heat and protease treatments show that the epitope recognized by the MAb Bb1, Db15 were a glycoprotein.

Cedar-apple rust fungi had been collected at 36 sites throughout the country from 1984 to 2001 and deposited at the Herbarium of Korea Forest Research Institute (HKFRI). We conducted the morphological examination on the dried specimens by light and scanning electron microscopy and as results six Gymnosporangium species were identified. Three species, G. asiaticum, G. clavaritforme and G. yamadae, were previously described in Korea, while the other three species, G. cornutum, G. globosum, and G. japonicum were new to Korea. Here we present the detailed morphological descriptions, distribution, host ranges and keys to species in both aecial and telial stages of each species. Some morphological characteristics related with telial formation on trees were newly identified; witches brooms for G. asiaticum, small galls for G. yamadae and telial formations on trunk for G. japonicun Geographically G. asiaticum and G. yamadae distributed widely throughout Korea, while the others were collected only at the limited locations. Eight Juniperus species as telial hosts and fifteen Rosaceous plants as aecial hosts were confirmed to be new in Korea.

Aecial host ranges of four Gymnosporangium species causing cedar-apple rust diseases, G. asiaticum, G. cornutum, 5. japonicum and G. yamadae, were investigated through artificial inoculation. Thirteen species of nine genera among Rosaceous plants, which have been reported as social hosts in Korea, were inoculated with fresh teliospores spores in early days of May of 2000 and of 2001, respectively. In the results, we re-confirmed that there was highly specific relationship between the rust species and aecial hosts and report new aecial hosts of four Gymnosporangium species. Teliospores of G. cornutum collected from Juniperus rigida successively produced spermogonia and aecia only on Sorbus alntifolia, the first report on host alteration of G. cornutum in Korea. Positive responses by teliospores of G. japonicum from J. chinenis of Suwon and from J. chinenis var. horizontalis of Jeju island were obtained only on P. villosa. Crataegus pinnatifida was confirmed as a new aecial host of G. viatium. Until this time, G. ymadae was believed to have Malus as the aecial host. However, teliospores of G. yamadae collected from J. chinensis var. kaizuka successively formed spermogonia and aecia on the leaves of Chaenomeles lagenaria, C. sinensis, Pyrus pyrtifolia var, culta, P. ussuriensis, Malus pumila and M. sileboldii. The date for maturation of spermogonia and aecia, and symptom development varied according to the rust fungi and aecial host plants, respectively.

The plant-pathogenic species S. scabies, S. acidiscabies, and S. turgidiscabies cause the scab disease of potato and produce the phytotoxins, thaxtomins. necl, a gene conferring a necrogenic phenotype, is involved in pathogenicity and physically linked to the thaxtomin A biosynthetic genes. Identification of the pathogenic strains of Streptomyces from soil was performed through the polymerase chain reaction by using specific pathogenicity primer sets derived from the necl gene sequences of Streptomyces smbies. The DNA was extracted from soil using a bead-beating machine and modifications of the FastPrep system. The DNA was suitable for direct use in the PCR. The PCR products showed the bands of approximately 460 bp. This methods can be very usuful in identifying species responsible for scab diseases and studying on the ecology of plant-pathogenic Streptomyces spp.

A technique for detection of Xanthomonas axonopodis pv. citri, a causal bacterium of canker on Unshiu orange fruits, was developed using bacteriophage. Procedure for the detection was designed on the basis of the previous reports that one group(CPI) of X. axonopodis pv. citri bacteriophage and corresponding two Iysotypes distributed in Korea. First, fruit surface was washed with sterile distilled water and pellet was obtained from centrifugation. The pellet was resuspended in Wakimoto's potato semi-synthetic broth medium and divided equally into two parts. One part was heated in boiling water to kill bacterial cells. Bacteriophages(CP$_1$) were respectively added into two parts and 0.1 ml from each part was mixed with soft agar medium. After incubation for 18 hrs at 25$^{\circ}C$, the causal bacterium of canker was determined based on plaques formed on the medium. This procedure can be effectively used for detection of living bacterial pathogen on fruit surfaces of Unshiu orange.

To characterize plasmids in Xanthomonu axonopodis pv. glycines, we isolated plasmids pAG1 from the strain AG1 and pXAG81 and PXAG82 from the strain Bra, respectively, and sequenced three plasmids. The size of plasmids, pAG1, pXAG81, and pXAG82 was 15,149-base pairs (bp), 26,727-bp, and 1,496-bp, respectively Fifteen and twenty six possible open reading frames (ORFs) were present in pAG1 and pXAG81, respectively. Only one ORF homologous to a rep gene of Xylella fastidiosa was present in pXAG82. pAG1 contained genes homologous to avrBs3, tnpA, tnpR, repA, htrA, three parA genes, M.XmaI, R.XmaI, and six hypothetical proteins. pXAG81 contained genes homologous to avrBs3, tnpA, tnpR, repA, htrA, two parA genes, pemI, pemK, mobA, mobB, mobC, mobD, mobE, trwB, traF, traH, ISxac2, and eleven hypothetical proteins. Based on DNA sequence analysis, we presume that pXAG81 is a conjugal plasmid. Interestingly, we found 0.5-kb truncated avirulence gene similar to aurXacE3 on the right border of avrBs3 homolgs of pAG1 and pXAG81. Two hundred twenty five isolates were analyzed to find aurBS3 or tra gene homologs by Southern hybridization. The numbers of avrBs3 homolog varied from 3 in AG1 to 8 in AG166. Two hundred seventeen isolates appeared to can conjugative plasmids (pXAG81 type), and thirty eight isolates appeared to carry non-conjugative plamids (pAGl type). This indicated that aurBs3 gene homologs might be spread by conjugation in X. axonopodis pv. glycines.

Bacterial grain rot by Burkholderia gluae cause severe damage in seedling and grain of rice after heading season. This seed-borne pathogen play a role as first infection agent that could be cause disease following cropping season. Until now the direct isolation of the bacteria has some trouble by interference of other bacteria existed inside seed. This study established convenient identification method as simple isolation with KB medium from seed showing symptom and using PCR identification. By this isolation method, B. glumae was isolated from 40 to 50％ in brown rice and inner hull, however, there were saprophytic bacteria and fungi outer hull. In PCR identification with Ogf4 and Ogr3 primer to these 25 isolates, the amplified products were presented in all of the collections but not in 10 saprophytic germs. The isolation rate was constant to 3 months stored seeds. This result provide a rapid and convenient isolation and identification of B. glumae.

In order to diagnose and differentiate jujube witches' broom (JWB) phytoplasma rapidly, oligonucleotide primer pair, 16Sr(V) F/R, for polymerase chain reactions (PCRs) was designed on the basis of 165 rRNA sequences of JWB phytoplasma. The PCR employing phytoplasma universal primer pair P1/P7 consistently amplified DNA in all tested phytoplasma isolates. But no phytoplasma DNA was detected in healthy jujube seedlings. The nested PCR, the primer pair 16S(V) F/R, about 460 bp fragment, amplified DNA in all tested JWB and related phytoplasmas including LiWB phytoplasma of the 165 rRNA group V, but no DNA amplification was detected from other phytoplasma strains such as group 16SrI (Aster yellows) and group 16SrⅩII (Stolbur group) phytoplasmas in which mulberry dwarf phytoplasma and chrysanthemum witches broom phytoplasma are belonged to, respectively The same results were obtained from both Korean- and Chinese-isolates of JWB. Nested-PCR using phytoplasma universal primer pair P1/P7 and 16S rRNA group V specific primer pair 16S(V) F/R could detect group V phytoplasma rapidly and easily, in particular JWB phytoplasma.

A pepper strain of Cucumber mosaic virus (Pf-CMV) induces a mild chlorotic spot symptom in zucchini squash at 9 days post-inoculation (dpi), wile Fny strain of CMV causes severe mosaic and stunting symptom at 4 dpi in this host. Pseudorecombinants were constructed between the two strains, and assessments of symptom severity were indicated that both RNA2 and RNA3 were responsible for both mildness and the slow appearance of symptom elicited by Pf-CMV in zucchini squash. With various RNA2 and RNA3 chimeras between two strains of CMV, the genetic symptom determinants of phenotype of Pf-CMV were mapped to Tyr residue at positions amino acid 267 in 2a protein and at positions amino acid 168 in 3a movement protein (MP). Chimeras changed the sequences (both changed Tyr to lie) in the codons of both amino acid 168 of 3a MP and amino acid 267 of 2a protein were resulted in the high RNA accumulation, severity of symptom, and the rapid systemic spread, suggesting that 2a replicase as well as MP is involved in virus movement. The RNA accumulation pattern of all pseudorecombinants and chimeras are identical in protoplast of zucchini squash, indicating the virus movement is responsible for the phenotypes of two CMV strains rather than virus replication.

The commercial cultivars of red pepper were screened against Tobacco mosaic virus (TMV), Pepper mild mottle virus (PMMoV) and Pepper mottle virus (PepMoV) by seedling test. Tn single infection of TMV or PMMoV, mosaic symptom was produced on the cultivars of 'Cheongyang'and 'Wangshilgun'. However, in cultivars of 'Manilla'and 'Bugang', symptoms were not occurred. In single infection of PepMoV, symptoms of mottle and malformation were produced on the tested cultivars of 'Manilla', 'Bugang', 'Cheongyang'and 'Wangshilgun' In the cultivars of 'Cheongyang'and 'Wangshilgun', synergistic symptoms of stunt and lethal death were induced by mixed infections in the two combinations of TMV+PepMoV and PMMoV+PepMoV. However, in cultivars of 'Manilla'and 'Bugang', synergistic symptom was not occurred as mottle which was milder than that of single infection. Cells were single infected with TMV and PMMoV the cultivars of 'Cheongyang'and 'Wangshilgun', respectively, had typical ultrastructures of tobamovirus as the stacked-band structure and multiple spiral aggregate (SA). Ultrastructures of cell and tissues infected with PepMoV on the cultivars of 'Cheongyang', 'Wangshilgun', 'Manilla'and 'Bugang', the potyvirus inclusions of pinwhills, scrolls, lamminated aggregates and amorphous inclusion were observed. Infected cells with a combination of TMV+PepMoV and PMMoV+PepMoV, the virus particles and inclusions of the two different viruses were found only mixed infection in the same cytoplasm and the amounts of viruses in mixed infections were abundant than in single infection. The angled-layer aggregates (ALA) was observed in the cells infected mixedly with TMV and PepMoV

Potato mop-top virus(PMTV) was identified from Solanum tuberosum cv. Gawon showing bright chlorotic mottle symptom in Namwon, Korea. Samples were collected green-house in February, 2003. Electron microscopic examination of negatively stained preparation revealed that PMTV were rigid-rod shaped particles about 100-150, 250-300 nm x 18-20 nm in length. In ultrathin sections of leaf tissue from diseased potato plants, cluster of viruses particles were observed in the cytoplasm. TAS-ELISA determined that the virus was serologically related to PMTV. PMTV produced double ring necrotic local lesion in inoculated leaf of Chenopodium amaranticolor in incubated at 15$^{\circ}C$. The PMTV could be detected with RT-PCR using PMTV detectable primer set designed to amplify about 540 bp of the partial CP gene of PMTV

Grapevine leafroll-associated 1 virus (GLRaV-1) and Grapevine leafroll-associated 3 virus (GLRaV-3), member of the genus Ampelovirus, are important viral disease of grapevine in the world. these viruses transmitted only dicotyledonous host by vectors such as mealybugs and there is no suitable herbaceous host for virus. The diseased leaves turn yellowish or reddish depending on cultivars and viruses. Viruses are existed at low concentration and ununiformly distribution in grapevine. Using small-scale double-stranded RNA (dsRNA) extraction method, reverse transcription and polymerase chain reaction (RT-PCR) product of 1Kb long which encoded of coat protein (CP) gene for both viruses was successfully amplified with a specific primers. The RT-PCR product was cloned into the plasmid vector and its nucleotide sequences were determined from selected recombinant cDNA clones. Sequence analysis revealed that the CP of GLRaV-1 consisted of 969 nucleotide, which encoded 323 amino acid residues and CP of GLRaV-3 consisted of 942 nucleotide, which encoded 314 amino acid residues. The CP of GLRaV-1 and GLRaV-3 has 93.8％ and 98.7％ amino acid sequence identities, respectively.

Tissues from woody plant contain higher amount of phenolic compounds and polysaccharides, which give inhibitory effects on reverse transcriptase and/or Taq ploymerase. The common multiple-step protocols using several additives to inhibit polyphenoic compounds during nucleic acid extraction are time consuming and laborious. Sodium sulfite (Na$_2$SO$_3$) was used as inhibitor of polyphenolic oxidases in extraction buffer and compare it's effect between commercial RNA extraction kit and small-scale double-stranded RNA (dsRNA) extraction by RT-PCR. During nucleic acid extraction procedure, addition of 0.5％-1.5％ (w/v) sodium sulfite to Iysis buffer or STE buffer resulted in lighter color change than extracts without sodium sulfite and improve the RT-PCR detection. When commercial RNA extraction kit used, optimal concentration of sodium sulfite were variable according to the host plant. However, using dsRNA as RT-PCR template, 1.5％ sodium sulfite in STE buffer improves the detection of both viruses and unspecific amplifications were reduced significantly, Furthermore, when viruses existed at low titers in host plant, small-scale dsRNA extractions were very reliable.

Ornithogalum showing mosaic symptoms were collected from the isolated field of National Plant Quarantine Service in Sengrimmyon of Kyungnam province. Electron microscopic examination of negatively strained preparation was filamentous particle of 740nm in lenght. Indirect-ELISA determined that the virus was serologically related to potyvirus. A single major protein band of Mr 30,000 was observed after sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Indicator plant test showed masaic, necrotic local lesion and sunken areas in leaves of Nicotiana clevelandii and Tetragonia expansa, while the others of indicator plants did not infect. An enzyme-aided purification protocal was used, which eliminated a highly viscous mucilage from extracts of the Omithogalum. Total RNA extracted from infected Omithogalum leaves were amplified of 411b.p fragment in reverse transcription (RT)-PCR when primers specilic for the coat protein gene. An isolate of Omithogalum mosaic virus (OrMV) of the genus Potyvirus was identified as the casual agent of the disease on the basis of electron microscopic, biological and serological reaction.

An Isometric virus was isolated from Paprika (Capsicum annuum var. glossum) showing necrosis spot and malformation on the fruit and the leaves, respectively, at yecheon in Korea. The virus could infect locally on Chenopodium amaranticolr, C. quinoa, Petunia x hybrida and Nicotiana glutinosa, but could not infect on Gomphrena globosa and Physalis floridana. The virus could infect systemically on red pepper and Lycopersicon esculentum. Datura stramonium, N. cleuarandii, N. rustim and N. tabacum cvs. were produced necrosis or necrotic ring spot lesions on the inoculated leaves and mosaic, vein necrosis or lethal death on the upper leaves. The virus was not related serologically to cucumber mosaic virus (CMV). In RT-PCR assay, it could not detected with specific primers of CMV and BBWV-II. The virions contain one molecule of genomic RNA, Which was approximately 3.8Kb and the coat protein (CP) of the purified virion migrated as a single band with molecular wight of about 29KDa in SDS-PAGE.

Among over-wintering small brown planthoppers, population of the rice stripe virus (RSV)-viruliferous insects was surveyed throughout the country in late April of 2003 by using DAS-ELISA. Averaged population of the RSV-viruliferous insects in this year was 2.1％, which was lower than that of last year of 3.7％. However, the insect population in Seoul, Incheon and Kyeonggi areas were relatively high showing 6.7％, 6.2％ and 2.6％, respectively. Based on the survey results, it was expected that overall occurrence of RSV on rice could be decreased in this year, except certain areas. Ovarial transmission rate of RSV by the insects on diseased rice samples collected from 10 areas ranged from 22.2％ to 77.8％. Among 35 graminous weed species collected from rice fields in Ganghwa and Kimpo in 2002 and 2003, common reed and formosens were found to be infected by RSV. The result indicates that those weeds are potential alternative natural hosts of the RSV Further studies on ecological and pathological impacts of the alternative natural host of RSV are being processed.

Forty six isolates of bean common mosaic virus (BCMV) collected from azuki bean, mungbean, kidney bean, cowpea, broad bean and peanut were classified into three groups based on biological, serological, cytopathological, and molecular characteristics. Group I induced vein-banding symptoms in cowpea which was similar to those produced by the BCMV-cowpea strain. Group II caused mosaic symptoms in azuki bean but not in peanut and tobacco. Since this character was different from that of previously described BCMV strain, group II may not belong to BCMV GroupIII induced vein-clearing symptoms in azuki bean, kidney bean and peanut, which are typical symptoms for BCMV-peanut stripe virus strain. Virus inclusion patterns of BCMV groups were similar to those of Potyvirus subdivision III with the scroll, pinwheel and long laminated inclusions. However, the inclusions of laminated aggregates were never observed in mungbean isolates. Multiple alignment as well as cluster dendrograms of 3'noncoding region (3'-NCR) and a part of coat protein gene (CP) suggested that group I belongs to the BCMV-cowpea strain, group II to the BCMV-azuki bean strain, and group III to the BCMV-peanut stripe virus strain. Since molecular phylogenesis of BCMV based on nucleotides of 3'-NCR and coat protein differed from the grouping based on virulence differentiation, and BCMV groups are more closely related to each other with the same host origin, other characteristics of those strains are under investigation.

A flexuous rod-shaped virus was isolated from Cucurbita pepo leaves showing green mosaic and puckering symptoms at Anseong, Korea. Based on the biological tests, electron microscopy, and reverse transcription-polymerase chain reaction (RT-PCR), the isolate was identified as Papaya ringspot virus type Watermelon (PRSV-W). In the biological test, host range of PRSV-W was limited in the families Cucurbitaceae and Chenopodiaceae. Most susceptible cucurbit species, such as Cucurmis lanatus, Cucurmis sativus, Cucurbita pepo, and Citrullus lanatus, responded to mechanical inoculation by PRSV-W that induce green mosaic, malformation, puckering, and narrow laminae. The local lesion symptoms were produced on the inoculated leaves of Chenopodium maranticolor and C. quinoa PRSV specific primers which amplifies the part of the coat protein (CP) genes, generated a 648 bp product from 6 isolates of PRSV-W, but no amplification had been detected in other viruses including CMV, CGMMV, KGMMV, ZYMV and WMV. In electron microscopy, PRSV particles were flexuous, approximately 780 nm in length and 12 nm in width. PRSV-W is one of the worldwide viruses which has the great economic importance in cucumber, melon, squash, watermelon, and other cultivated cucurbits with ZYMV and WMV. This is the first report of PRSV-W on cucurbits in Korea.

Hop stunt viroid(HSVd) is a plant pathogen which infect a number of hosts such as grapevine, Citrus and Prunus plants. Sequence variants of HSVd have been divided into three types(i. grapevine and hop, ii. citrus, iii. plum, peach, apricot and almond). Purified RNAs from plum trees were used for the synthesis of cDNA with reverse transcription and amplified by polymerase chain reaction. Cloned cDNAs were sequenced and two different consensus sequence variants were detected. A neighbor-joining analysis was carried out on the sequence variants together with 62 previously described variants of HSVd from hop, plum and other species. Sequence variants from plum trees cultivated in Korea were clustered in HSVd-plum subtype and not in HSVd-hop subtype which were two Korean isolates belongs. These relationship between sequence variants from plum and two Korean isolates in HSVd-hop type supports the other origin for hop stunt disease.

A rapid and sensitive assay for the specific detection of plant viroids using reverse transcription-polymerase chain reaction(RT-PCR) has been developed already. The nested RT-PCR assay cloud be applied for the detection of apple scar skin viroid(ASSVd) from young leaves and other tissues. ASSVd has central conserved region(CCR), terminal left(T$\sub$L/) and terminal right(T$\sub$R/) domain. Primers were designed from these regions. Primer sets were successfully applicable for the amplification of full length or partial region of ASSVd by nested RT-PCR. Nested RT-PCR assay was more sensitive and accurate method to detect ASSVd from young trees during the early time of apple cultivation.

A Korean isolate of Pepper mild mottle virus (PMMoV-Kr) was isolated from a diseased pepper crop in Chunchon, Korea. The isolate was biologically purified on Nicoticaa tabacum cv. Xanthi-nc by successive single local transfer steps, and propagated on N. tabacum cv. Samsun. PMMoV-Kr could systemically infect on N. glauca, N. benthmiana, N. occidentalis and Lycopersicon esculentum, which is typical of known isolates of PMMoV. PMMoV-Kr belongs to the pathotype P1,2 based on pepper-tobamoviral indicator experiments; Capsicn chinone harboring L3 gene revealed resistant (necrotic local lesion on inoculated leaf, HR) whereas L+, L1 and L2 pepper plants expressed susceptible reactions of mosaic systemic symptoms for the isolate. To confirm the pathology and delineate symptom determinant of the isolate, full-length cDNAs of PMMoV-Kr were amplified by RT-PCR with a primer set corresponding to the 5'- and 3'-ends of PMMoV. The RT-PCR molecules amplified from genome RNA of the isolate was cloned into the pUC18 vector. Full-length cDNA clones constructed under the control of the T7 RNA promoter could be successfully transcribed to produce in vitro transcript RNA. Infectivity of the capped transcripts and its progeny virus was verified by Western blot and RT-PCR analyses.

Attenuated viruses can protect their hosts against challenge to their related viruses. Increasing evidence shows that mutations of the tobamoviral 126/183 kDa protein play a major role in the viral attenuation and contribute to the cross protection mechanism. In this study, four mutants of Pepper mild mottle virus (PMMoV) have been constructed by mutagenesis; two mutants, pTPpoly348 and pTPpoly762, were substituted in the middle of replicase gene, and the others, pTPL3D:: $\Delta$6207 and pTPL3D:: $\Delta$6219, were deletion mutants made by deleting some parts of pseudoknot structures of the 3' noncoding region (NCR) of the virus. Progeny viruses generated from the four mutants were infectious on N. benthamiana plants with symptomless or mild mosaic symptom. Replication efficiency and viral product accumulations of four mutants were assessed by Northern and Western blot analyses on BY-2 protoplast cells. Accumulation of CP for the pTPL3D:: $\Delta$6207 and pTPL3D:: $\Delta$6219 were lower than that of other mutants and wild type virus. These data suggest that the 3'-NCR mutations contribute to the viral gene expression in host tissues, while mutants of replicase gene rather govern the symptom expression.

This study was performed to investigate the incidence of Hosta uirus X (HVX), a Potexvirus, from cultivated hosta ornamental plants in Korea and to ascertain seed transmission of the virus from infected parent plant to progeny ones for breeding program of hosta plants. Infection rate of HVX in cultivated hostas was 25.6 ％ (11 out of 43 collected samples contained HVX) based on Western blot and RT-PCR detection methods. Most of HVX-infected hostas showed visible systemic leaf symptoms (mosaic, mottle, curling, stunting or combinations). Variability of HVX was confirmed by sequences of coat protein gene of individual isolates from different hostas. HVX was seed-transmitted on Hosta 'Blue Cadet'. The virus was detected from seeds, and sprouts and seedlings from the virus-contaminated seed sources. Over 7.5 ％ of seeds were HVX-contaminated surveyed in this study, Our data suggest that HVX can be transmitted by seed source, and indexing of the virus should be done for breeding program of Hosta.

This study was conducted to designing conserved regions of molecules for virus-derived resistance to transgenic Phlaenopsis orchids to protect against two major orchid viruses, Cymbidum mosaic virus (CymMV) and Odontoglossum ringspot virus (ORSV). Infected leaf samples of Phalaenopsis were randomly screened by the RT-PCR with specific primers to both of viruses. RT-PCR products of the viruses were cloned and their nucleotide sequences were determined. Multiple alignments of coat protein (CP) genes of the viruses revealed that over the 88 ％ and 94 ％ identities with CymMV and ORSV, respectively, were observed. These data can be useful for selection of highly conserved regions of CP gene of the viruses for transgenic orchid experiments.

To investigate occurrence and variability of satsuma mandarin ( Citrus unshiu)-infecting viruses in Jeju island, several sets of diagnostic RT-PCR primers were designed and applied to samples collected randomly. Each primers set used in this survey was designed to detect Satsuma dwarf virus (SDV, Sadwavirus) and Citrus mosaic virus (CiMV) which is reclassified as an isolate of SDV (SDV-CiMV, Saduavirus). RT-PCR methods could detect SDV-CiMV and CTV from leaf . samples of unshui citrus. CTV was the prevalent and SDV-CiMV was not common in Jeju island. RT-PCR product of SDV-CiMV-JJl2 were cloned and sequenced. Sequence of the isolate revealed that it was 96.9 ％ identical to SDV-CiMV-Jp isolate at the nucleotide level. SDV-CiMV-JJl2 was propagated on Physalis floridana and sequencing of entire sequences of genome is in progress. Variability of SDV in Jeju island was confirmed by sequence comparisons and restriction mapping analysis.

Transgenic Nicotiana benthmiana plants harboring and expressing coat protein (CP) gene of Zucchini green mottle mosaic virus (ZGMMV) were generated for both virus-resistant screening and complementation analysis of related viruses and environmental safety assessment (SA) of living modified organism (LMO) purposes. Transformation of leaf disc of N. benthamiana was performed using Agrobacterium-mediated method and the pZGCPPGA748 containing the ZGMMV CP and NPTII genes. Two kinds of transgenic homozygous groups, virus-resistant and -susceptible lines, were obtained by screening of challenging homologous virus for T1 generations. Complementation of CP-deficient related virus was analyzed using the susceptible line of ZGMMV. These two pathologically different lines can be useful for host-virus interactions and LMO environmental SA.

Two Korean isolates of Alfalfa mosaic virus (AHV-AZ, AMV-KR) were isolated from azuki bean and potato plants, respectively, and their pathologies were confirmed on some susceptible host plants including pepper, tobacco and red bean plants. Full length cDNAs to RNA1, RNA2 and RNA3 of the two Korean strains were amplified using the long-template reverse transcription (RT)-polymerase chain reaction (PCR) method. RT-PCR products covering entire regions for the three AMV genome RNAs were cloned. RNA transcripts were synthesized in vitro from each clones using T7 RNA polymerase and infectivity test was peformed in 9 reassortment sets of transcripts. All the combinations of reassorted transcripts were found to be infectious when inoculated onto Nicotiana benthamiana plants, and were not distinguishable to those of wild types. The full-length cDNA clones that were confirmed infectious were sequenced their nucleotide sequences. We will discuss sequence analysis of the two Korean isolates of AMV genomic RNA3 and compare reported foreign isolates of AMV.

Freesia, a member of the Iridaceae family, has fragant, tubular shaped flowers and is very popular ornamental plants in the world. Diseased freesia plants showing systemic leaf streak mosaic symptoms were collected from a cultivated farm in Kyonggi province, Korea in 2003, and its causal agents were investigated. Two viruses, Cucumber mosaic virus (Fr-CMV) and a potyvirus, were identified from the leaf tissues of the diseased freesia based on sequence analysis and host range tests. CMV-Fr could infect systemically on Chenopodium quinoa, C. amaranticolor, N. glutinosa, and N. benthamiana, and this biological property is distinguishable from ordinary strains of CMV. A filamentous potyvirus-shaped virus could not infect general indicator plants by mechanical inoculation. Single RT-PCR products was successfully amplified with a set of degenerate primers specific to the Potyvirus genus and total nucleic acids from the infected tissues, and was cloned into the pGEMT-Easy vector. Nucleotide sequences confirmed it belongs to the Potyvirus genus with either a new species or an isolate of Freesia mosaic virus (no information is available for the FrMV). This is the first report of FrMV in Korea and more characterizations of the two viruses are in progress.

Complete nucleotide sequence of genome RNA of a Korean isolate of Pepper mottle virus (PepMoV-Vb) from field-collected diseased paprika (Capsicum annuum var grossum) was determined in this study. Symptoms of isolates of PepMoV were divided largely into two groups, vein banding (Vb) and vein clearing (Vc) patterns. PepMoV-Vb RNA consists of 9,640 nucleotides excluding the poly(A) tail. A single open reading frame was identified beginning at nucleotide position 169 encoding a polyprotein of 3024 amino acids which is typical of the Potyvirus genus. The complete nucleotide sequence and coding regions of PepMoV-Vb were compared to that of 11 potyviruses within the genus Potyvirus. The overall nucleotide sequence identity was 94.7 and 94.1％ identical to PepMoV-C and PepMoV-FL, respectively. Full-length cDNAs of PepMoV-Vbl were synthesized from purified viral RNAs by RT-PCR and their genome structure was analysed by RFLP analysis. This is the first report on complete nucleotide sequence of PepMoV isolated from paprika in Korea.

Natural virus infection of cultivated Daphe odora plants showing chlorosis and stunting was observed and their causal agent was investigated. An isolate of isometic virus was purified from infected leaf tissues, and it could infect systemic severe mosaic on Chenopodium quinoa and C. amaranticolor. cDNA library was generated from partially purified viral RNAs and oligo dT primer-pSPORTl system, and recombinant clones were selected and their inserts were sequenced randomly. Nucleotide sequences of the virus were analyzed by BLAST, and it was closely related to members of subgroup B in the genus Nepovirus. The sequence analysis suggest that the virus was identified as an isolate of Cycas necrotic stunt virus (CNSV) because it was 89.7 ％ and 94.7 ％ identical to known CNSV for the CP and 3' noncoding region, respecitively. RT-PCR was performed to screen disease incidence of CNSV in Daphe plants, and five out of 10 plants (50 ％) were infected by CNSV This is the first sequence information of CNSV from Daphe plants.

Cucumber fruit mottle mosaic tobamovirus (CFMMV) causes severe mosaic symptoms with yellow mottling on leaves and fruits, and occasionally severe wilting of cucumber plants. No genetic source of resistance against this virus has been identified. The genes coding for the coat protein or the putative 54-kDa replicase were cloned into binary vectors under control of the SVBV promoter. Agrobacterium-mediated transformation was peformed on cotyledon explants of a parthenocarpic cucumber cultivar with superior competence for transformation. R1 seedlings were evaluated for resistance to CFMMV infection by lack of symptom expression, back inoculation on an alternative host and ELISA. From a total of 14 replicase-containing R1 lines, 8 exhibited immunity, while only 3 resistant lines were found among a total of 9 CP-containing lines. Line 144 homozygous for the 54-kDa replicase was selected for further resistance analysis. Line 144 was immune to CFMMV infection by mechanical and graft inoculation, or by root infection following planting in CFMMV-contaminated soil. Additionally, line 144 showed delay of symptom appearance following infection by other cucurbit-infecting tobamoviruses. Infection of line 144 plants with various potyviruses and cucumber mosaic cucumovirus did not break the resistance to CFMMV. The mechanism of resistance of line 144 appears to be RNA-mediated, however the means is apparently different from the gene silencing phenomenon. Homozygote line 144 cucumber as rootstock demonstrated for the first time protection of a non-transformed scion from soil inoculation with a soil borne pathogen, CFMMV.

A lily strain of Cucumber mosaic virus (LK-CMV) was not able to systemically infect zucchini squash (Cucurbita pepo), while Fny strain of CMV (Fny-CMV) caused systemic mosaic and stunting symptom at 4 days post-inoculation on the same host species. The pathogenicity of LK-CMV in zucchini squash was investigated by reassortments of genomic RNAs of LK-CMV and Fny-CMV for infection, as well as by pseudorecombinants generated from biologically active transcripts of CDNA clones of LK-CMV and Fny-CMV, respectively. The assessments of pathogenicity for LK-CMV indicated that RNA2 of LK-CMV was responsible for systemic infection in zucchini squash. In the protoplast of zucchini squash, the RNA accumulations of all constructed pseudorecombinants were indistinguishable and LK-CMV replication was slightly lower than that of Fny-CMV, suggesting that the inability of LK-CMV to infect squash plants was responsible for the poor efficiency of virus movement, rather than the reduction of replication function.

Occurrences of virus diseases on paprika ( Capsicum annuum var. grossum) were surveyed in Joennam province from 1999 to 2003 and the collected samples showing virus-like symptoms were tested using ELISA. Virus diseases appeared 4.5％, 17.5％, and 4.9％ in 2000, 2002, and 2003, respectively. As the results of investigation of the seasonal incidence with the growing stages of plant, virus was not occurred at seedling stage and was slightly from the planting time to the first harvesting time, but was dramatically increased at the second harvesting time. Virus diseases were more severe on the vinyl house than on the green house. Pepper mild mottle virus (PMMoV) was severely occurred in 2000 but not after that year. Comparing the virus species, Pepper mottle virus (PepMoV) was 35.9％, Broad bean wilt virus (BBWV) was 14.1％, and Cucumber mosaic virus (CMV) was 10.9％ in 2002, and 76.0％, 11.1％, and 2.4％ in 2003, respectively.

A single-step multiplex reverse transcription polymerase chain reaction (RT-PCR) assay was developed for the simultaneous detection of five cucurbit-infecting viruses： cucumber mosaic virus (CMV), watermelon mosaic virus 2 (WMV2), zucchini yellow mosaic virus (ZYMV), cucumber green mottle mosaic virus (CGMMV), and kyuri green mottle mosaic virus (KGMMV). The multiplex RT-PCR provides a simple and rapid method for detecting various viruses in cucurbit plants, which will help diagnose many cucurbit plants at a time.

Carnation mottle carmovirus(CarMV) was isolated from Lilium spp. in Korea. This isolate, CarMV, was done bioassay, which plants were Dianthus caryophyllus, Gomphrena globosa, Chenopodium amaranticolor, Dianths chinensis. CarMV was propagated on the leaves of Chenopodium amaranticolor with the crude-sap inoculation method and purified by Mossops method(1976). We produced antiserum against CarMV and analyzed the antiserum specificity with ELISA, Gel diffusion method, and Rapid Immunofilter Paper Assay (RIPA). From these results of the assay, RIPA method was simple and rapid for CarMV detection. We have established successfully the CarMV detection system. CarMV coat protein gene was amplified by RT-PCR with specific primers and sequencing analysis was done.

To analyze the genome of Cucumber mosaic virus(CMV) in pepper, we developed a new extraction method for double-stranded RNA(dsRNA). To isolate the dsRNA, 0.1g of pepper leaves homogenized with 1ml of 5${\times}$EXB extraction buffer[0.5M glycin, 0.5M NaCl, 5mM EDTA(pH9.0/NaOH), 10％ Sodium N-lauryl salcosinate(NLS), 10％ Sodium dodecylsulfate(SDS)] and purified with the 1/4 volume of phenol： chloroform： isoamylalcohol(25：24：1). dsRNAs from the aqueous phase was precipitated with isopropanol. This procedure was able to detect a minimal amount of dsRNA from CMV infected plant tissue and to distinguish different CMV satellite RNAs by polyacrylamide gel electrophoresis(PAGE). Moreover, this method can be applied CMV infected in pepper or Rice dwarf virus (RDV) infected rice.

Chinese yam(D. opposita Thunb. cv. Jang-Ma) plants showing necrotic mosaic symptom were collected from their growing fields in Andong, Korea. Electron microscoptic examination of negatively stained preparations showed filamentous particles of 660nm in length. The virues purified partially were used to isolate Viral RNA as template for RT-PCR to amplify the CP gene with ChYNMV specific and oligo dT primers. Amino acids sequeces revealed that the viruses shared 99.3％ similarity with ChYNMV(AB044386) which was known as the member of macluravirus. So the viruses from Chinese yam(D. opposita Thunb. cv. Jang-Ma) plants were identified as ChYNMV. Comparing the amino aced sequences of ChYNMV strains with other macluraviruses such as CdMV, NLV and MacMV revealed that N-terminal was the most variable region and conserved regions were present within the genus Macluravirus.

One fungus was isolated from lesions on imported chinese cabbage leaves in process of quarantine inspection from China. The fungus was identified as Mycosphaerlla brassicicola, based on morphology of perithecia, asci, ascospore, and curtural characteristics. In Korea, this fungus has been quarantine fungus, and not yet report to occur. Perithecia of the fungus were globose, dark brown with apical papilate ostioles. The size was 90-100 ${\times}$ 130-135$\mu\textrm{m}$. Asci were bitunicate, 8-spored and 38-43 ${\times}$ 15-l9$\mu\textrm{m}$. Ascospore were irregularly biseriate, hyaline, cylindrical, 2-celled, and rounded at the ends. Optium growth temperature of the fungus was at 20$^{\circ}C$ on PDA but did rarely grow over 24$^{\circ}C$. Colony on PDA was of black aerial mycelia.