• Journal of Applied Biological Chemistry
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• v.61 no.4
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• pp.405-410
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• 2018

Pseudomonas tolaasii strain No. 6264 has been isolated from mushroom tissue and identified as one of the major pathogen causing brown blotch disease. It secretes peptide toxins, known as tolaasin and its analogue peptides. P. tolaasii 6264 has been used as a typical pathogenic strain to study the brown blotch disease for last 20 years after confirming its blotch-forming ability, hemolytic activity, and white line formation. In this study, the characteristics of P. tolaasii 6264 strain were analyzed and compared according to storage period. Strains of P. tolaasii 6264 stored annually since 2012 were cultured and their pathogenic characters were analyzed. When the 16S rRNA sequences were compared, all strains were divided into two groups. Pathogenic characters including hemolytic activity, blotch-forming ability, and white line test were also investigated. The strains, P. tolaasii 6264-15-2 and P. tolaasii 6264-17, had all three activities; however, the rest of stored strains showed only blotch-forming ability losing other pathogenic characters. Tolaasin peptides were purified from the bacterial cultures and analyzed by mass spectrometry. The strains, P. tolaasii 6264-15-2 and P. tolaasii 6264-17, secreted Tol I (1987 Da), Tol II (1943 Da), and its analogues (1973 Da, 2005 Da) while some of these peptides were not found in the media cultured other strains. These results indicate that the pathogenicity of P. tolaasii could be varied during the storage period.

• Korean Journal Plant Pathology
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• v.10 no.3
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• pp.197-210
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• 1994

This experiment was carried out to study the cause of degeneration and rot of cultivated mushroom. Among 597 bacterial isolates derived from the rots of Button mushroom (Agaricus bisporus), Oyster mushroom (Pleurotus ostreatus) and Oak mushroom (Lentinus edodes) collected from markets of 5 cities (Seoul, Suwon, Taegu, Pohang and Pusan) in Korea (1991~1993), 111 bacterial isolates (18.5%) were proved as pathogenic bacteria. These pathogenic bacteria causing bacterial rots of cultivated mushrooms were identified as Pseudomonas tolasii, P. agarici, and Eriwinia sp., and the main causal bacteria were P. tolaasii. P. fluorescens and Klebsiella plenticola were confirmed as saprophytic non-pathogenic bacteria. One hundred fifty nine isolates (Group No. 39) of the 486 saprophytic bacterial isolates were classified as P. fluorescens, and this species was most often found rot area of cultivated mushrooms. P. tolaasii, the causal organism of bacterial blotch, was classified into two groups; One group can be differentiated from the other by the formation of white precipitation band by white line reacting organisms of Pseudomonas Agar F media. P. tolaasii attacked the cultivated mushrooms relatively well at lower incubation temperature such as 5$^{\circ}C$, but P. agarici rarely attack at below 1$0^{\circ}C$. The temperature for the infection commercial cultivated mushrooms by P. agarici was higher than that of P. tolaasii. Optimum temperature for the infection of mushrooms by P. tolaasii and P. agarici were 2$0^{\circ}C$ and $25^{\circ}C$, respectively.

• Proceedings of the Korean Biophysical Society Conference
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• 1998.06a
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• pp.34-34
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• 1998

Tolaasin is a channel forming bacterial toxin produced by Pseudomonas tolaasii and causes a brown blotch disease on cultivated oyster mushrooms. When tolaasin molecules form channels in the membranes of mushroom cells, they destroy cellular membrane structure, known as 'colloid osmotic lysis'. In order to understand the molecular mechanisms forming membrane channels by tolaasin molecules, we have investigated the electrophysiological characteristics of tolaasin-induced channels in lipid bilayer.(omitted)

• Proceedings of the Korean Biophysical Society Conference
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• 1999.06a
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• pp.47-47
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• 1999

Tolaasin is a bacterial paptide toxin which is produced by Pseudomonas tolaasii. It forms pores in the cellular membranes, causing the brown blotch disease on the cultivated oyster mushroom. Previously, we showed that tolaasin-induced pore formation required the multimerization of tolaasin molecules. In order to measure the ionic effect on the tolaasin multimerization, the time course of tolaasin-induced hemolysis was measured in the presence of various cations and anions.(omitted)

• Korean Journal of Organic Agriculture
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• v.24 no.3
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• pp.571-582
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• 2016

Bacterial fruit blotch (BFB) is one of most important diseases in Cucurbitaceae due to infection of Acidovorax citrulli, causing huge economic losses damage worldwide. This seedborn disease spread rapidly at period of high temperature and humidity. The eco-friendly farming is getting popular. So far there was no effective agent to control BFB in eco-friendly farming. Therefore, effect of the material based on chinese nut-gall extract with antibacterial activity against BFB to was tested against A. citrulli. Different hosts showed various symptoms of BFB. Liquid formulation among exhibited most effective anti-bacterial activity on A. citrulli. Pot experiment in greenhouse showed the potential as an control agent of BFB in cucurbits. The treatment of material based on chinese nut-gall extract showed the positive effect on survival of the watermelon seedling and on the length of the cucumber seedling treated with A. citrulli. We cautiously conclude that the material based on chinese nut-gall extract used in this study may be good agents against major diseases of cucurbits in the future even though it is require to be tested with more study on field test.

• Korean Journal of Organic Agriculture
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• v.23 no.2
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• pp.323-334
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• 2015

This study was conducted to develop environment-friendly agricultural products with anti-microbial activity against Acidovorax avenae subsp. citrulli as a pathogen of bacterial fruit blotch in cucurbit. Schlechtendalia chinensis was extracted by MeOH and solvent fraction. The hexane fraction, which showed highest value of anti-microbial activity, was analyzed by GC-MS. Each mass spectra, corresponding to each peak of chromatogram, was compared to MS database of Wiley library. As a result, myristic acid, palmitic acid and 3-n-pentadecylphenol were identified as maine compounds showing antimicrobial activity against A. avenae subsp. citrulli. Bioassay using commercial myristic acid, palmitic acid and 3-n-pentadecylphenol to test for the anti-microbial activity conformed the anti-microbial activity of potential active compounds, and myristic acid and 3-n-pentadecylphenol showed strong activity. In conclusion, myristic acid and 3-n-pentadecylphenol identified from S. chinensis were anti-microbial chemicals.

• Korean Journal Plant Pathology
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• v.11 no.4
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• pp.353-360
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• 1995

Tests were performed on 232 bacterial strains (71 strains of Pseudomonas tolaasii and 161 white line reacting organisms, WLRO) isolated from cultivated mushrooms. As results, P. tolaasii was divided into 5 groups on the basis of the phenotypical characteristics for the strains, and group 3 was the major one including 48 (62%) out of the total 71 strains. WLRO were classified into 23 groups, and group 10 was the major group (65 strains, 30% of the total WLRO tested). A white line was well formed at 22$^{\circ}C$ and at 4 mm distance between P. tolaasii and WLRO colonies in their dual culture on Pseudomonas agar F medium within 36-hr incubation, but not formed at $25^{\circ}C$ even for 72-hr incubation. The morphological, cultural and biological properties of P. tolaasii group 3, and the main group of WLRO, group 10, were different only in the light of pathogenicity. Also group 2 of P. tolaasii had the characteristics similar to group 24 of WLRO which was pathogenic to cultivated mushrooms, suggesting the P. tollaasiii and WLRO strains may be the same species although their white line forming ability and pathogenicity were more or less different from one another. Subculture of the strains in P. tolaasii group 1 induced the variation of their pathogenicity, white line forming ability and utilization of sodium benzoate and sodium tartrate, and thus it can be inferred that they were converted to strains having the characteristics of group 3 or WLRO groups.

• The Plant Pathology Journal
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• v.33 no.2
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• pp.170-183
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• 2017

Bacterial fruit blotch (BFB), which is caused by Acidovorax citrulli, is a serious threat to watermelon growers around the world. The present study was conducted to screen effective rhizobacterial isolates against 35 different A. citrulli isolates and determine their efficacy on BFB and growth parameters of watermelon. Two rhizobacterial isolates viz. Paenibacillus polymyxa (SN-22), Sinomonas atrocyanea (NSB-27) showed high inhibitory activity in the preliminary screening and were further evaluated for their effect on BFB and growth parameters of three different watermelon varieties under greenhouse conditions. The greenhouse experiment result revealed that SN-22 and NSB-27 significantly reduced BFB and had significant stimulatory effect on total chlorophyll content, plant height, total fresh weight and total dry weight compared to uninoculated plants across the tested three watermelon varieties. Analysis of the 16S ribosomal RNA (rRNA) sequences revealed that strains SN-22 belong to P. polymyxa and NSB-27 to S. atrocyanea with the bootstrap value of 99% and 98%, respectively. The isolates SN-22 and NSB-27 were tested for antagonistic and PGP traits. The result showed that the tested isolates produced siderophore, hydrolytic enzymes (protease and cellulose), chitinase, starch hydrolytic enzymes and they showed phosphate as well as zinc solubilizing capacity. This is the first report of P. polymyxa (SN-22) and S. atrocyanea (NSB-27) as biocontrol-plant growth promoting rhizobacteria on watermelon.

• Journal of Mushroom
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• v.9 no.3
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• pp.91-95
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• 2011

Pleurotus ostreatus, the oyster mushroom, is one of the most important edible mushrooms. It is especially susceptible to bacterial blotch disease, which is caused by Pseudomonas tolaasii. In order to develop bacterial blotch disease-resistant transgenic mushroom, NUECIN cDNA, a gene for an antibacterial peptide cloned from Bombyx mori, was overexpressed in Pleurotus ostreatus. NUECIN cDNA was fused to the ${\beta}$-TUBULIN promoter of oyster mushroom and co-transformed with the pTRura3-2 vector into the uracil auxotrophic mutant strain. Twelve transformants containing the NUECIN gene were identified by genomic PCR and Southern blot analysis. NUECIN gene expression was confirmed by Northern blot analysis. Three transformants showed the transcriptional expression of the gene. However, we could not detect expression of the protein in the transformants. This study showed the possibility of transgenic mushroom development for disease resistance.

• The Plant Pathology Journal
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• v.37 no.1
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• pp.36-46
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• 2021

Acidovorax citrulli (Ac) is the causal agent of bacterial fruit blotch (BFB) in watermelon, a disease that poses a serious threat to watermelon production. Because of the lack of resistant cultivars against BFB, virulence factors or mechanisms need to be elucidated to control the disease. Glycerol-3-phosphate dehydrogenase is the enzyme involved in glycerol production from glucose during glycolysis. In this study, we report the functions of a putative glycerol-3-phosphate dehydrogenase in Ac (GlpdAc) using comparative proteomic analysis and phenotypic observation. A glpdAc knockout mutant, AcΔglpdAc(EV), lost virulence against watermelon in two pathogenicity tests. The putative 3D structure and amino acid sequence of GlpdAc showed high similarity with glycerol-3-phosphate dehydrogenases from other bacteria. Comparative proteomic analysis revealed that many proteins related to various metabolic pathways, including carbohydrate metabolism, were affected by GlpdAc. Although AcΔglpdAc(EV) could not use glucose as a sole carbon source, it showed growth in the presence of glycerol, indicating that GlpdAc is involved in glycolysis. AcΔglpdAc(EV) also displayed higher cell-to-cell aggregation than the wild-type bacteria, and tolerance to osmotic stress and ciprofloxacin was reduced and enhanced in the mutant, respectively. These results indicate that GlpdAc is involved in glycerol metabolism and other mechanisms, including virulence, demonstrating that the protein has pleiotropic effects. Our study expands the understanding of the functions of proteins associated with virulence in Ac.